TECHNICAL REPORT EPA/CE-81-1

              ENVIRONMENTAL PROTECTION AGENCY/
                     CORPS OF ENGINEERS
               TECHNICAL COMMITTEE ON CRITERIA
                FOR DREDGED AND FILL MATERIAL
PROCEDURES FOR HANDLING AND CHEMICAL ANALYSIS
           OF  SEDIMENT AND  WATER SAMPLES
                           May 1981
                         PUBLISHED BY
                   ENVIRONMENTAL LABORATORY
          U. S. ARMY ENGINEER WATERWAYS EXPERIMENT STATION
                      VICKSBURG, MISSISSIPPI

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vvEPA
              ENVIRONMENTAL PROTECTION AGENCY/
                      CORPS OF ENGINEERS
               TECHNICAL COMMITTEE ON CRITERIA
                 FOR DREDGED AND FILL MATERIAL
PROCEDURES FOR HANDLING AND GHEMICAL ANALYSIS
           OF SEDIMENT AND WATER SAMPLES
                              by

                       Russell H. Plumb, Jr.
                      Great Lakes Laboratory
                  State University College at Buffalo
                      1300 Elmwood Avenue
                     Buffalo, New York  14222
                           May 1981
        Prepared for U.S. Environmental Protection Agency/Corps of
              Engineers Technical Committee on Criteria
                    for Dredged and Fill Material

                    Under Contract EPA-480557201 0

                  Monitored by  Large Lakes Laboratory
                U.S. Environmental Protection Agency
                        9311 Groh Road
                    Grosse Me, Michigan 48138

                 published by Environmental Laboratory
           U.S. Army Engineer Waterways Experiment Station
             P.O. Box 631, Vicksburg, Mississippi  39180

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    Destroy this report when no longer needed.  Do not return
                      it to the originator.
The findings in this report are not to be construed as an official
     Department of the Army position unless so designated.
                by other authorized documents.
       The contents of this report are not to be used for
       advertising, publication, or promotional purposes.
       Citation of trade  names  does not constitute an
       official  endorsement or approval of the use  of
                  such commercial products.

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                                   PREFACE

            This project was supported by Grant EPA-lj.805572010 between
the Environmental Protection Agency and the Research Foundation of the
State University of New York.  Funding for this project was equally
shared between the U. S. Environmental Protection Agency and the U. S.
Army Engineer Waterways Experiment Station.  The objective of the effort
was to prepare a procedures manual that will contain summaries and
descriptions of the tests, sample collection and preservation procedures,
analytical procedures, and calculations required for the evaluation of
Section ^OH permits as specified in Public Law 92-500.
            This work was conducted during the period March 1978 -
March 1980 by the Great Lakes Laboratory (GLL), State University College
of New York at Buffalo, Buffalo, New York.  The investigation was con-
ducted by Dr. Russell H. Plumb, Jr., Associate Director, GLL.  The study
was under the general supervision of Dr. Robert A. Sweeney, Director,
GLL.
            The contract was monitored by Mr. Jim Westhoff and
Dr. Robert M. Engler of the Environmental Laboratory (EL), U. S.  Army
Engineer Waterways Experiment Station, Vicksburg, Mississippi; and
Dr. Michael D. Mullin, U. S. Environmental Protection Agency, Grosse
lie Laboratory, Grosse lie, Michigan.  Directors of WES during the con-
duct of this study and preparation of this manual were COL J. L.  Cannon,
CE, and COL N. P. Conover, CE.  Technical Director was Mr. F. R.  Brown.

            This report should be cited as follows:
            Plumb, R. H., Jr.  198l.  "Procedure for Handling
            and Chemical Analysis of Sediment and Water Sam-
            ples," Technical Report EPA/CE-81-1, prepared
            by Great Lakes Laboratory, State University Col-
            lege at Buffalo, Buffalo, N. Y., for the U. S.
            Environmental Protection Agency/Corps of Engi-
            neers Technical Committee on Criteria for Dredged
            and Fill Material.  Published by the U. S. Army
            Engineer Waterways Experiment Station, CE, Vicks-
            burg, Miss.

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                           TABLE OF CONTENTS
Preface
List of Tables
List of Figures
Conversion Factors
INTRODUCTION

SECTION l:  PROJECT MANAGEMENT GUIDANCE
            Project Definition
            Test Selection
               Testing procedures
               Chemical analyses
            Sampling Considerations
               Representative sampling
               Sampling techniques selection
               Sample preservation
            Quality Control
            Additional Considerations
            Summary
            References

SECTION 2:  FIELD/LABORATORY GUIDANCE
            Method of Sample Collection
               Water samplers
               Sediment samplers
            Sample Collection
            Sample Handling
            Sample Preservation
                                                                    Page
  i
  x
xii
 xv
  1

1-1
1-2
1-3
1-3
1-5
1-5
1-6
1-19
1-22
1-2U
1-25
1-26
1-27

2-1
2-1
2-2
2-3
2-U
2-11
2-15
                                11

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    Total Solids Procedure for Water Samples
    Method 1:   Gravimetric
    Total Solids Procedure for Sediment Samples
    Method 1:   Gravimetric
  Volatile Solids Determination                       3-59
  Specific Gravity                                    3-6l
    Procedure for Sediment Samples                    3-6l

Inorganic Analysis                                    3-6H
  Carton, Total Organic and Inorganic                 3-65
    Procedure for Water Samples                       3-69
    Method 1:   Infrared Analysis                      3-69
    Procedure for Sediment Samples-                    3-73
    Method 1:   Sample Ignition                        3-73
    Method 2:   Differential Combustion                3-7^
  Metals (Al,  Cd, Ca, Cr, Cu, Fe, Pb, Mg,
          Mn,  Mo, Ni, Zn).                             3-77
    Procedure for Water Samples (All metals
      except As, Hg, and Se)                          3-80
    Method 1:   Direct Flame Atomic Absorption,
               Total Metals                           3-80
    Method 2:   Direct Flame Atomic Absorption,
               Soluble Metals                         3-85
    Method 3:   Graphite Furnace Atomic
               Absorption                             3-89
    Method k:   Chelation-Extraction Atomic
               Absorption                             3-93
    Procedure for Sediment Samples (All metals
      except As, Hg, and Se).                          3-96
    Method 1:   Direct Flame Atomic Absorption,
               Total Metals                           3-96
  Metals (Arsenic)                                    3-110
    Procedure for Water Samples                       3-110
    Method 1:   Arsine Generation                      3-110
    Method 2:   Graphite Furnace                       3-
                       iv

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            Quality Control                                        2-20
               Objectives                                          2-20
               Work load                                           2-22
               Present limitations                                 2-25
            Types of Chemical Tests                                2-25
               Elutriate test                                      2-28
               Sediment fractionation                              2-32
               Bulk analysis                                       2-38
            Summary                                                2-kk
            References                                             2-U5

SECTION 3.  ANALYTICAL METHODS                                     3-1
            Physical Analysis                                      3-19
               Cation Exchange Capacity                            3-20
                  Procedures for Sediment Samples                  3-23
                  Method 1:  Agitation, Filtration                 3-23
                  Method 2:  Centrifugation                        3-25
               Particle Size                                       3-28
                  Procedures for Sediment Samples                  3-33
                  Method 1:  Sieving and Electronic Particle
                             Counters                              3-33
                  Method 2:  Sieving and Pipet Analysis            3-39
               PH                                                  3-U8
                  Procedures for Water Samples                     3-1+9
                  Method 1:  Glass Electrode                       3-^9
                  Procedures for Sediment Samples                  3-51
                  Method 1:  Glass Electrode                       3-51
               Oxidation Reduction Potential                       3-52
                  Procedures for Water and Sediment Samples        3-52
                  Method 1:  Platinum Electrode                    3-52
               Total Solids and Volatile Solids                    3-5^
                                   111

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  Procedure for Sediment Samples                 3-116
  Method 1:  Arsine Generation                   3-116
Metals (Mercury)                                 3-118
  Procedure for Water Samples                    3-118
  Method 1:  Cold Vapor Technique                3-118
  Procedure for Sediment Samples                 3-I2k
  Method 1:  Cold Vapor Technique                3-12U
Metals (Selenium)                                3-127
  Procedure for Water Samples                    3-127
  Method 1:  Hydride Generation                  3-127
  Procedures for Sediment Samples                3-131
  Method 1:  Digestion/Flameless Atomic
             Absorption                          3-131
  Method 2:  Hydride Generation                  3-133
Nitrogen (Ammonia, Nitrate, Nitrite, Total
  Kjeldahl, Organic)                             3-138
Nitrogen (Ammonia)
  Procedures for Water Samples
  Method 1:  Colorimetric, Automated Phenate
  Method 2:  Colorimetric, Automated
             0-Tolidine
  Method 3:  Colorimetric or Titrimetric,
             Manual                              3-1^8
  Procedures for Sediment Samples                3-15^-
  Method 1:  Potassium Chloride Extraction       3-15^
  Method 2:  Distillation                        3-155
  Method 3:  Distilled Water Extraction          3-157
Nitrogen (Nitrate),                               3-159
  Procedures for Water Samples                   3-159
  Method 1:  Colorimetric, Manual, Brucine
             Sulfate                             3-159
  Method 2:  Colorimetric, Automated, Cadmium
             Reduction                           3-163

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                                                  Page
  Method 3:  Colorimetric, Automated,
             Hydrazine Reduction                  3-l68
  Method 4:  Colorimetric, Manual, Cadmium
             Reduction                            3-174
Nitrogen (Nitrite)                                3-179
  Procedures for Water Samples                    3-179
  Method 1:  Colorimetric, Automated, Cadmium
             Reduction                            3-179
  Method 2:  Colorimetric, Automated,
             Hydrazine Reduction                  3-179
  Method 3:  Colorimetric, Manual, Cadmium
             Reduction                            3-179
  Method 4:  Colorimetric, Manual                 3-179
  Procedure for Sediment Samples                  3-183
Nitrogen (Total Kjeldahl)                         3-185
  Procedures for Water Samples                    3-185
  Method 1:  Colorimetric, Semiautomated with.
             Block Digestor                       3-185
  Method 2:  Manual Colorimetric, Titrimetric     3-190
  Method 3:  Colorimetric, Automated Phenate      3-195
  Procedures for Sediment Samples                 3-201
  Method 1:  Kjeldahl Digestion                   3-201
  Method 2:  Block Digestion                      3-202
Nitrogen (Organic)                                3-205
Phosphates (Soluble Reactive, Total, Organic)     3-207
Phosphates (Soluble Reactive)                     3-212
  Procedures for Water Samples                    3-212
  Method 1:  Ascorbic Acid, .Manual                3-212
  Method 2:  Ascorbic Acid, Automated             3-213
  Method 3:  Stannous Chloride, Manual            3-214
  Method 4:  Stannous Chloride, Automated         3-218
  Method 5:  Vanadomolybdophosphoric Acid,
             Manual                               3-221
  Procedure for Sediment Samples                  3-223
                  vi

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  Phosphates (Total!
    Procedures for Water Samples
    Procedures for Sediment Samples
  Phosphates (Organic)                                3-230
    Procedures for Water Samples                      3-230
    Procedures for Sediment Samples                   3-232
    Method 1:  Acid Hydrolysis                        3-232
    Method 2:  Acid Extraction                        3-233
  Sulfides                                            3-236
    Procedures for Water Samples                      3-238
    Method 1:  Methylene Blue, Colorimetric           3-238
    Method 2:  Iodine Titrimetric                     3-2^0
    Procedure for Sediment Samples                    3-2^3
    Method 1:  Distillation, Methylene Blue,
               Colorimetric                           3-2^3

Organic Analysis                                      3-2^8
  Carbamates                                          3-2^9
    Procedure for Water Samples                       3-2^9
    Method 1:  Methylene Chloride Extraction          3-2^9
    Procedure for Sediment Samples                    3-255
    Method 1:  Methylene Chloride Extraction          3-255
  Chlorinated Phenoxy Acid Herbicides                 3-260
    Procedures for Water Samples                      3-262
    Method 1:  Chloroform Extraction                  3-262
    Method 2:  Ethyl Ether Extraction                 3-265
    Procedure for Sediment Samples                    3-271
    Method 1:  Acetone-Hexane Extraction              3-271
  Oil and Grease                                      3-278
    Procedure for Water Samples                       3-278
    Method 1:  Freon Extraction                       3-278
                      VI1

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  Procedure for Sediment Samples                   3-28H
  Method 1:  Freon Extraction                      3-28*1
Chlorinated Hydrocarbons                           3-289
  Procedures for Water Samples                     3-291
  Method 1:  Benzene Extraction                    3-291
  Method 2:  Methylene Chloride/Hexane
             Extraction                            3-298
  Procedures for Sediment Samples                  3-307
  Method 1:  Acetone/Hexane Extraction             3-307
  Method 2:  Acetonitrile Extraction               3-313
Organophosphorous Pesticides                       3-319
  Procedure for Water Samples                      3-321
  Method 1:  Hexane, Chloroform, Benzene
             Extraction                            3-321
  Procedure for Sediment Samples                   3-332
  Method 1:  Hexane Extraction                     3-332
Polynuclear Aromatic Hydrocarbons                  3-337
  Procedure for Water Samples                      3-337
  Method 1:  Dichloromethane Extraction/Gas
             Chromatography                        3-337
  Procedures for Sediment Samples                  3-3^1
  Method 1:  Methanol Extraction/UV Analysis       3-3^1
  Method 2:  Ethanol Extraction/
             UV Spectrophotometry                  3-3UU
Phenolics                                          3-3U8
  Procedures for Water Samples                     3-3^8
  Method 1:  Distillation, H-aminoantipyrine
             Colorimetric                          3-3^8
  Method 2:  Distillation, MBTH Colorimetric       3-352
  Procedures for Sediment Samples                  3-355
  Method 1:  Distillation, H-aminoantipyrine
             Colorimetric                          3-355
                  Vlll

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     Method 2:  MBTH Colorimetric
Miscellaneous Analysis
  Chlorine Demand
    Procedures for Water Samples
    Procedures for Sediment Samples
  Biochemical Oxygen Demand
    Procedures for Water Samples
    Procedures for Sediment Samples
  Chemical Oxygen Demand
    Procedures for Water Samples
    Method 1:  Low Level, 5 to 50 mg/
    Method 2:  High Level, 50 to 800-mg/
    Procedure for Sediment Samples
  Sediment Oxygen Demand
    Procedure for Sediment Samples
    Method 1:  In Situ
    Method 2:  Laboratory
                      IX

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                            LIST OF TABLES
No.                                                                Page
1-1   Analytical Costs by Parameter                               1-12
1-2   Performance Requirements, Number of Samples Per Day
      Per Man                                                     1-15
2-1   Operational Evaluation of Grab Samplers                     2-5
2-2   Operational Suitability of Corers and Grab Samplers         2-9
2-3   Recommended Method of Sample Storage as a Function of
      Bulk Sediment Analyses to be Performed                      2-lU
2-U   Recommended Water Preservation Techniques                   2-17
2-5   Ideal Control Activities for Documenting the Validity of
      Laboratory Data                                             2-23
3-1   Acceptable Test Procedures                                  3-6
3-2   Comparison of Scales Used to Report Particle-Size Results   3-30
3-3   Comparison of Particle-Size Distribution Analytical
      Methods                                                     3-30
3-1*   Data Tabulation for Sand-Size Fractions                     3-37
3-5   Data Tabulation for Coulter Counter Results for Silt-
      and Clay-Sized Fractions                                    3-38
3-6   Maximum Sieve Loads on 8-in.-diam Sieves for 1-phi
      Intervals                                                   3-^2
3-7   Sampling Time Intervals for Pipet Analysis                  3-^5
3-8   Typical Particle-Size Distribution Data Sheet               3-^6
3-9   Recommended Atomic Absorption Spectrophotometer Instru-
      ment Settings for Metal Analysis                            3-87
3-10  Graphite Furnace Operating Conditions for Selected Metals   3-91
3-11  HN03-HC1 Digestion                                          3-98
3-12  HN03 Digestion                                              3-100
3-13  HN03-H202 Digestion                                         3-101
3-1^  Muffle Furnace Ignition                                     3-103
3-15  Low Temperature Ashing                                      3-10U
3-16  HF-HClO^HNOa Digestion                                     3-105
3-17  HF-HN03-HC1 Digestion                                       3-106
3-l8  Operating Characteristics for TKN AutoAnalyzer Manifold     3-189
3-19  Some N-methylcarbamates and Related Compounds               3-250
3-20  Retention Times of Various Organochlorine Pesticides
      Relative to Aldrin                                          3-297

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No.                                                               Page
3-21  Persistence of Chlorinated Hydrocarbon Pesticides in
      River Water in Terms of Percentage Recovery                 3-306
3-22  Composition of Organophosphorous Nanogram Standard          3-323
3-23  Detection Limit for 1^ Organophosphorous Pesticides in
      1-& Water Samples                                           3-326
3-2U  Retention Time and Peak Height Data for Organophosphorous
      Pesticides                                                  3-329
3-25  Retention Time for Organophosphorous Compounds on OV-101/
      OV-210 at 210C                                             3-330
                                  XI

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                            LIST OF FIGURES
No.                                                               Page
1-1   Average concentration as a function of dredge penetra-
      tion                                                        1-21
2-1   Comparative methods of sediment sample handling             2-13
2-2   Elemental partitioning procedure for sediment characteri-
      zation                                                      2-39
2-3   Sediment sample splitting for bulk analysis                 2-U2
3-1   Schematic of generalized sample handling procedure          3-3
3-2   Handling and storage procedures for cation exchange
      capacity samples                                            3-22
3-3   Handling and storage procedures for particle-size samples   3-32
3-^   Particle-size procedure using sieving/pipet analysis        3-^1
3-5   Handling and storage procedures for total and volatile
      solids samples                                              3-55
3-6   Handling and storage procedures- for total inorganic
      carton samples                                              3-66
3-7   Handling and storage procedures for total organic carbon
      samples                                                     3-67
3-8   Handling and storage procedures for metals samples          3-79
3-9   Hydride generator for arsenic and selenium analysis         3-111
3-10  Handling and storage procedures for mercury samples         3-119
3-11  Schematic cold vapor apparatus for mercury                  3-120
3-12  Handling and storage of samples for ammonia analysis        3-1^1
3-13  AAI manifold for phenate determination of ammonia           3-1^5
3-1^  AAII cartridge for phenate determination of ammonia
3-15  AA manifold for the 0-tolidine determination of ammonia
3-l6  Handling and storage of samples for nitrate analysis        3-160
3-17  Copper cadmium reduction column                             3-l65
3-l8  AAI manifold for nitrate determination following cadmium
      reduction                                                   3-l67
3-19  AAII cartridge for the determination of nitrate following
      cadmium reduction                                           3-169
3-20  AAI manifold for the determination of nitrate following
      hydrazine reduction                                         3-172
3-21  AAII cartridge for the determination of nitrate follow-
      ing hydrazine reduction                                     3-173
                                  XII

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No.                                                               Page
3-22  Nitrate reduction column                                    3-175
3-23  Handling and storage of samples for nitrite analysis        3-l80
3-2k  Handling and storage of samples for TKN analysis            3-186
3-25  AAII cartridge for ammonia determinations with TKN
      digests                                                     3-187
3-26  AAI manifold 1 for the determination of total Kjeldahl
      nitrogen                                                    3-198
3-27  MI manifold 2 for the determination of total Kjeldahl
      nitrogen                                                    3-199
3-28  Continuous digester for the automated determination of
      total Kjeldahl nitrogen                                     3-200
3-29  Handling and storage of samples for orthophosphate
      analysis                                                    3-209
3-30  Handling and storage of samples for organic phosphate
      analysis                                                    3-210
3-31  Handling and storage of samples for total phosphate
      analysis                                                    3-211
3-32  MI manifold for the ascorbic acid determination of
      phosphorus                                                  3-215
3-33  Mil cartridge for the ascorbic acid determination of
      phosphorus                                                  3-216
3-3^  Stannous chloride manifold for the determination of
      phosphorus, low level                                       3-219
3-35  Stannous chloride manifold for the determination of
      phosphorus, high level                                      3-220
3-36  Handling and storage of samples for sulfide analysis        3-237
3-37  Handling and storage of samples for carbamate analysis      3-251
3-38  Handling and storage of samples for chlorophenoxy acetic
      acid analysis                                               3-26l
3-39  Handling and storage of samples for oil and grease analy-
      sis                                                         3-279
3-UO  Handling and storage of samples for chlorinated hydro-
      carbon analysis                                             3-290
3-^1  Handling and storage of samples for organophosphate
      analysis                                                    3-320
3-^2  Handling and storage of samples for polynuclear aromatic
      hydrocarbon analysis                                        3-338
3-1*3  Handling and storage of samples for phenol analysis         3-3^9
                                 XI11

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No.                                                               Page
3-^  Handling and storage of samples for chlorine demand
      analysis                                                    3-363
3-^5  Handling and storage of samples for biological oxygen
      demand analysis                                             3-375
3-^6  Handling and storage of samples for chemical oxygen
      demand analysis                                             3-386
3-^7  In situ sediment oxygen demand chamber                      3-398
3-^8  Laboratory sediment oxygen demand chamber                   3-^00
                                  xiv

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              CONVERSION FACTORS, U. S. CUSTOMARY TO METRIC  (Si)
                             UNITS OF MEASUREMENT
U. S. customary units of measurement used in this report can "be converted to
metric (.Si) units as follows:
	Multiply	                   By                      To Obtain
feet                                  0.3CA8                meters
gallons (U. S. liquid)                3-785^12              cubic decimeters
inches                               25.^                   millimeters
miles (U. 3. statute)                 1.6093^7              kilometers
pounds (mass)                         0.^53592^             kilograms
square miles                          2.589998              square killometers
                                   xv

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                             INTRODUCTION

      1.  The purpose of this handbook is to provide state-of-the-art
guidance on the subjects of sampling, preservation, and analysis of
dredged and fill material.  This need developed as a result of the
promulgation and implementation of Section UoU(b) of Public Law (PL)
92-500 (Federal Water Pollution Control Act Amendments of 1972) which
required the ecological evaluation of proposed dredging and filling
operations as they may impact navigable waters of the United States.
It is expected, therefore, that this manual will receive wider usage
as an aid in the regulatory process rather than a research tool.
      2.  The initial guidance for implementing Section UoH(b) was
released in 1976. *  The guidance presented in this handbook should be
viewed as second-generation Interim Guidance in the continuing process
of procedure development, refinement, and evaluation.  Thus, it will
be intermediate between the initial Interim Guidance and analytical
compendiums such as Standard Methods,2  American Society of Testing and
Materials (ASTM) manuals,3 or Environmental Protection Agency (EPA)
manuals.1*  The major emphasis of this effort has been to provide
guidance on the subjects of sampling, sample handling, and sample
pretreatment.
      3.  Three approaches were used to obtain the information required
to prepare this manual.  They were:
          &_.  Review published literature on sediment sampling and
              analysis.
          b_.  Contact personnel at several laboratories that are active
              in the field of sediment investigation:
              (l)  U. S. Environmental Protection Agency (EPA) Research
                   Laboratories
              (2)  U. S. Army Engineer Waterways Experiment Station
                   (WES)
              (3)  Universities
          c_.  Contact personnel involved in the regulatory process,
              requesting suggested input:
              (1)  EPA Regional Offices
              (2)  U. S. Army Engineers District Offices

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This information was compiled and presented in one of three major sections:
          a_.  A discussion of rationale for project managers.
          b_.  A step-by-step protocol for sample handling and each test
              procedure.
          c_.  A listing of analytical techniques, including sample pre-
              treatment procedures.
      k.  The purpose of the first section is to point out to a project
director or project manager the types of trade-offs that have to be
considered in developing an acceptable sampling program.  Unfortunately,
it is not possible to give project-specific guidance in a manual such
as this.  However, if a project director is aware of the kind of infor-
mation provided by use of each piece of equipment or testing procedure,
and the present limitations of this information, he can then make a
decision to use the equipment and/or procedures that are most suited
to his particular project.
      5-  The second section of the handbook provides guidance to the
laboratory and field personnel that will be implementing the sampling
program.  This includes a discussion of the types of sampling equipment
to be used and when to use each type, a step-by-step description of the
three general chemical tests considered, along with the required method
of sample handling, and a general quality control program, beginning
with sample collection.  The three chemical tests that are described
are:
          a_.  A short-term water leaching test (the standard elutriate
              test).
          b_.  A strong acid digest or an organic solvent extract
              (bulk analysis).
          c_.  An elemental distribution test (sediment fractionation) .
      6.  The third section presents for laboratory personnel a series
of analytical techniques, including sample preparation, where required,
for hh parameters.  Since the purpose of this manual was not to develop
new methods, the methods are generally thos.e found in Standard Methods,
ASTM, and EPA manuals.  The listed procedures are considered most
appropriate for general use; it is recommended that they be utilized
when it is decided to analyze samples for that particular constituent.
However, the fact that UU procedures are included is not meant to imply

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that all tests should be run on all samples or that additional parameters
should not be considered where appropriate.

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                              References
1.  Environmental Effects Laboratory.   "Ecological Evaluation of Proposed
    Discharge of Dredged or Fill Material into Navigable Waters."
    Interim Guidance for Implementation of Section HoH(b)(i)  of Public
    Law 92-500 (Federal Water Pollution Control Acts Amendments of
    1972).  Environmental Effects Laboratory,  U.S. Army Engineer
    Waterways Experiment Station, Vicksburg, Mississippi.  Report
    D-76-17.  83 p. (1976).

2.  American Public Health Association.  Standard Methods for the
    Examination of Water and Waste Water Including Bottom Sediments,
    and Sludges.  lUth Edition:   APHA, New York, New York.  1193 p.
    (1975).

3.  American Society for Testing Materials.  "Part 31.  Water."
    American Society for Testing Materials, Philadelphia, Pennsylvania
    (1976).

h.  U. S. Environmental Protection Agency.  "Methods for Chemical
    Analysis of Water and Wastes."  Environmental Monitoring and
    Support Laboratory, U.S. EPA; Cincinnati,  Ohio (l979).

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                SECTION 1:  PROJECT MANAGEMENT GUIDANCE

          The initiation of a sediment, soil, and water sampling program
is a difficult task because of the large number of factors that must be
considered.  These factors include such technical decisions as:
          eu  Selection of sampling locations.
          b_.  Selection of sampling equipment.
          c_.  Number of samples to collect.
          d_.  Type of tests to be performed on the samples.
          e_.  Specific chemical analyses to be performed.
The purpose of this handbook is to provide state-of-the-art guidance,
particularly for personnel involved in regulatory programs such as
Section UoU(b) of PL 92-500, for sampling and chemical characterization
of contaminants associated vith dredged and fill materials.
          Two levels of guidance are provided to satisfy the purpose of
this handbook.  One level consists of a decision as to how to handle a
sample following collection or how to perform a specific chemical test
once a decision has been made to run the test.  This information is
directed to the field personnel or laboratory personnel that will be
processing the samples and is found in Section 2 and Section 3.  The
second level of guidance is directed towards project managers or other
administrative personnel that decide where to collect samples or what
tests are to be run.  This information is presented in the remainder
of this section.
          Any guidance provided to project managers, by necessity,
must be more subjective than that provided to laboratory and/or field
personnel.  For example, in designing a generalized sampling program,
it would be ideal to specify more samples and replicates rather than
fewer, the use of corers rather than grab samplers, and the running
of all tests and all chemical analyses.  Such an approach would
maximize the information obtained from a project area and result in a
broader environmental evaluation.  However, it is realized that there
will be times when real-world constraints such as manpower and/or
fiscal limitations will limit this approach.  At other times, project
goals may be so explicit that it is not necessary to run all tests.

                                 1-1

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Since factors such as fiscal limitations and project goals are site
specific and cannot be addressed in a general manual, the following
discussion will summarize the administrative decisions that must be
made in establishing a sampling program.  Therefore, a project manager
will be aware of how the information to be gained by including a
specific test benefits his program, and what information is sacrificed
by excluding a test from his program.

                          Project Definition

          An essential component of any field sampling program is a
preproject meeting with all concerned personnel.  The list of concerned
personnel in attendance should include representatives from management,
field operations, and laboratory operations.  The purpose of the meeting
should be to define the objective of the sampling program and to ensure
communication between participating groups.
          The function of conducting a sampling program and performing
specific tests on the samples is to gain information.  It should be
obvious that the amount of information gained will be directly propor-
tional to the number of samples collected and the number of tests
performed.  However, the information generated through a sampling
program must be directed at a specific need.  The purposes of defining
the objective(s) of a sampling program should be to clarify the infor-
mation needed and to match these needs with the specific tests that
supply the required information.
          The definition of a project should be more specific than
"an environmental assessment of a proposed dredge or fill material
disposal operation."  Although an environmental assessment may be the
overall objective, this objective would be considered a cumulative
effect.  Therefore, the objective of the sampling program should be
subdivided into specific tasks such as:
          a_.  Compare two or more sites in a project area.
          b_.  Quantitate the total amount of certain contaminants
              present.
          c_.  Determine the mobility of contaminants in dredged and/
                                 1-2

-------
              or fill material.
          cL  Determine the distribution of certain chemical contaminants
              in the sediments of a project area.
          e_.  Determine potential sediment toxicity.
          f_.  Determine the biological suitability of project site water.
          _.  Determine whether a local discharge has altered the water
              and/or sediments in the project area.
          h_.  Determine the sediment phase distribution of certain
              chemical contaminants in the sediments of a project area.
The more explicitly the goals of a project can be stated, the easier it
should be to select the tests to be run and the method of sample handling.

                            Test Selection

Testing procedures
          Another aspect of the planning meeting is to determine the
specific tests and analyses to be completed.  The selection of the type
of tests for water samples is generally limited to water quality tests
and bioassays.  These results can then be evaluated by contrasting the
results with established water quality criteria and interpreting the
bioassay results.
          Types of tests available for the analysis of sediments include:
          ji.  Bulk analysis.
          b_.  Standard elutriate test.
          c_.  Fractionation/extraction procedures.
          d_.  Physical analysis.
          e_.  Bioassay.
          f_.  Bioaccumulation.
The utility of any one sediment test for a particular project can only
be determined after the purpose of the study has been identified since
each test provides different information as indicated below:
          a^.  Bulk analysis provides an estimate of the total con-
              centration of a constituent in the sediment sample.
              The analytical.result will include the various sediment
              phases (interstitial water phase, exchangeable phase,
              residual phase, etc.) but is poorly related to the
              biological availability of the constituent'.  A beneficial
                                 1-3

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    aspect of this test is that storage and presentation
    problems are minimized since changes in the oxidation
    state generally do not affect total concentrations.
    Bulk analysis results are useful for calculating an
    inventory of the total amount of a constituent involved
    in a dredging or filling project.  However, a major
    limitation of the test is that the results are a poor
    indicator of the potential environmental effects of
    moving the material (as in a dredging operation) "because
    of the poor relationship between biological availability
    and total concentration.
b_.  The standard elutriate test provides an estimate of the
    mobility of chemical constituents from sediments to the
    water column.  This test has the advantage of being more
    environmentally interpretable since it measures "water-
    soluble" constituents, which are the basis of most water
    quality criteria.  The disadvantages associated with the
    elutriate test are the lack of understanding of the
    mixing process, which could influence interpretation;
    the fact that the test is of short duration and may not
    estimate long-term changes following disposal; and the
    fact that this test, like bulk analysis, does not address
    possible impacts on the benthic fauna.  In addition, the
    test requires a greater effort for storage and preservation
    of samples since oxidation-state changes can alter test
    results.
c_.  Fractionation procedures provide more detailed information
    on the distribution of chemical constituents within the
    sediments by subjecting the sample to a series of
    increasingly harsh extraction solutions.  It is possible
    that there may be a crude inverse relationship between the
    harshness of the extraction solution and the bioavailability
    of the constituents.  However, the full meaning of a given
    distribution is not understood.  Further limitations of
    this procedure are that the actual testing is more time-
    consuming and strict storage requirements are mandatory.
dL  Physical analysis can provide information on particle
    size, mineralogical characterization, color, and texture.
    Some of the results can provide an indication of the
    adequacy of the sampling program.  Storage and preser-
    vation requirements are generally minimal.
.  Bioassay procedures are beyond the scope and purpose of
    this manual.  However, their use may be required to
    evaluate the potential biological response to a partic-
    ular dredged material disposal or fill material disposal
    operation.1  Strict storage and preservation of samples,
    similar to that used for fractionation procedures or the
    elutriate test, are required.
                       l-U

-------
          f_.  Bioaccumulation studies are a subset of bioassay
              procedures.  Their use may be required to determine
              whether chemical contaminants of interest can be
              concentrated to undesirable levels by locally important
              organisms under in situ or controlled laboratory
              conditions.  Sample storage time should be kept to an
              absolute minimum as sample dehydration and/or tissue
              deterioration can affect calculated bioconcentration
              factors.
Chemical analyses
           Similarly, the specific chemical analyses to be performed
should be discussed so that sufficient sample is collected at each
location and proper methods of sample preservation and storage are
available.  The selection of chemical parameters to be analyzed should
be based on major point sources and contaminants of concern in the
project area.  Because of the site-specific variability of point
sources, no mandatory or minimum list of analyses can be recommended.
           Once a list of specific chemical analysis to be performed
has been finalized, the required sample containers and appropriate
preservation techniques are also determined.  This follows from the
fact that sample handling and preservation techniques are mandatory
for each chemical constituent.  While this may be the last decision
to be made regarding sample handling, it is important to realize that
it must be made prior to actual sample collection so that appropriate
materials are available at the time of collection.  The need for and
limitations of available preservation techniques are discussed later.

                        Sampling Considerations

           A critical aspect of any environmental evaluation is the
field phase during which the samples to be analyzed are collected.  The
importance of this component is underscored by the fact that the quality
of any such evaluation is only as good as the information gained through
sampling.  Thus, any errors incurred during sampling will manifest
themselves by limiting the accuracy and/or the appropriateness of the
study.
           The objective of such collections is to obtain samples from
                                1-5

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                                                                   2
a project area with the purpose of characterizing the area sampled.
Sample size should "be small enough to be conveniently handled and
transported and yet sufficient to meet the requirements for all planned
analyses.  The quality of the information obtained through the sampling
process is dependent upon:
           a_.  Collecting representative samples.2 '3'"*
           b_.  Using appropriate sampling techniques.
           c_.  Protecting the samples until they are analyzed (sample
               preservation). '
Ideally, each of the three factors will be fully understood for each
project.  In practice, however, this is not always the case.  There may
be times when fiscal, time, or other resource constraints will limit
the amount of information that can be gathered.  When this occurs, each
of these factors must be carefully considered in light of specific
project purposes when designing a sample collection program.
Representative sampling
           Several criteria have been established to define the
representative nature of a sample.  It is considered mandatory that:
           a_.  The project area being sampled is clearly defined.
           b_.  The sampling locations are randomly distributed.
           c_.  More than one sample is collected from each sampling
               location unless the sample variability has been pre-
               established.
           d_.  If sample variability is unknown, it may be necessary to
               conduct a preliminary survey of the project area to better
               define the final sampling program.
In defining the project area, it must be remembered that sediment
composition can vary in the vertical dimension as well as in the
areal dimensions.  Thus, samples should be collected over the entire
project depth unless the sediments are known to be homogeneous in the
vertical dimension.  The purpose of collecting random samples is to
define the range of chemical concentrations or characteristics that
may be found in the project area.  The easiest task in establishing
a sampling program is to locate the areas of maximum concentrations
that are generally located near major point sources and/or areas of
quiescent settling.  However, results from these sampling locations

                                 1-6

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are not representative of the range of concentrations  in the  project
area.  Therefore, additional sampling must be conducted throughout
the remainder of the project area.   The last two criteria (c  and d)
relate to the number of samples that should be collected.   In effect,
the number of samples required is inversely proportional to the  amount
of known information and proportional to the level of  confidence
that is desired in the results.
           In addressing the question of representativeness,  it  is
possible to define two populations: '   one population is the actual
composition of sediments in the project area; the second population
is the composition of the sediment samples obtained from the  project
area.  Ideally, these populations will be the same.  However, it is
necessary to be aware of the fact that differences may exist  between
these two populations because of bias in the sampling  program.   Factors
that can contribute to the bias are oversampling near  major point
sources and equipment limitation (i.e. extrapolating surface  grab
sample results to subsurface sediments).
           Any established field program should be sufficiently
flexible to allow changes based on field observations.  The hetero-
geneity of the water column can be readily assessed through the  use
of electrometric probes for conductivity, dissolved oxygen, and  pH.
If the vertical profiles for these parameters are uniform, water
column sampling probably can be minimized.  However, if stratification
is indicated, sampling should be conducted in each of  the defined
layers.  Sediments generally are more heterogeneous than the  water
column, but the variability is not as easy to compensate for  in  the
field.  Certain characteristics of the sediments, such as color  or
texture, will provide an indication of patchiness.  The greater  the
patchiness, the larger the number of samples that will be required to
define the project area.  Other valuable sources available to refine
a sampling program can be historical data and/or a preliminary sampling
survey at the site.
           Ultimately, it must be admitted that representativeness is
one of the most difficult concepts to reduce to operational practice.
                                 1-7

-------
The condition of representativeness is essential to most statistical
tests; but, despite its fundamental importance, the only positive
practice that can be advocated is to be avare of the possible intro-
duction of bias during sampling.
           Sampling site locations.  The EPA  has identified seven
factors that should be involved in sampling site selection.  These
factors are:
           a_.  Objectives of the study.
           b_.  Accessibility.
           c_.  Flows.
           d..  Mixing.
           .  Source locations.
           f_.  Available personnel and facilities.
           g_.  Other physical characteristics.
The actual sampling pattern to be used, by necessity, has to be site
dependent because major point sources, land use activities, hydro-
logic conditions, and sample variability fluctuate from area to area.
           The pattern should consider major point sources in the
project area and currents that could be critical to the sediment
distribution pattern.   Thus, primary station locations should be
located downstream from major point sources and in quiescent areas,
such as turning basins, side channels, and outside channel bends,
where fine-grained sediments are most likely to settle.  The intensity
of sampling around point sources will be influenced by project
purposes and available resources.  If the purpose is to identify and
locate point sources,  a higher sampling density will be needed.
Additionally, samples should be collected above and below the
discharge so the relative input and impact can be ascertained.  If
the purpose of the project is to characterize sediments and/or fill
material that will be affected by a dredging or filling operation,
a lower sample density, proportional to sample heterogeneity, should
suffice.
           Sampling in quiescent areas of harbors, rivers, and channels
is advocated because these areas are conducive to the settling of
                                1-8

-------
finer materials.  The importance of this size fraction is that the
concentration of many constituents is concentrated within the smaller
particle sizes.  For example, Helmke et al.7 and Forstner et al.8
have shown higher concentrations of metals in the <2 y size fraction.
Helmke et al.,7 in particular, have shown that this size fraction
delineates sources and dispersion much better than total sediment
analysis.
           Sampling patterns based on the above suggestions would  be
logical.  In addition, a knowledge of the point sources would provide
a basis for selecting the parameters for which analyses should be
completed.  A limitation with this approach is that the sample results
will be biased high.  That is, the primary sampling locations are
situated near point sources where concentrations would be expected
to be higher (or highest) and in quiescent areas with a higher
percentage of fine material (and a concurrently higher concentration
of associated contaminants).  It may be desirable under certain
circumstances (i.e. regulatory agencies may want to know maximum
concentrations) to collect such samples; but, as a consequence, the
samples cannot be considered representative of the concentration
distribution in the project area; the validity of extrapolating
conclusions from these samples to the entire project area is
questionable.  Therefore, in order to provide some degree of
representativeness, the primary sampling locations listed above
should be supplemented with random stations located throughout the
project area.  At the present time, no firm guidance can be given
on the number of additional sampling stations that should be
established.  However, it is suggested that the number of such
additional sampling stations be equal to or slightly greater than  the
number of sampling stations located in the vicinity of major point
sources to compensate for the high bias.
           An additional factor that should be included in establishing
a sampling program is the selection of a reference station and/or  a
control station.  Data from a reference or control station are required
for a site comparison, as outlined in the Federal Register.9  Sediments
                                 1-9

-------
in such areas are subject to the same heterogeneity as discussed

earlier.  To compensate for this variability, it is recommended that

reference area sampling be replicated.
           The following general guidance is offered as an aid in

establishing a sampling program:
           a_.  Sampling stations should be located downstream from
               major point sources in the project area.  These sources
               may be selected based on specific constituents in the
               effluent or the volume of the discharge.  It is usually
               possible to define these sources based on a knowledge
               of the activity in the area or a review of historical
               data for the site.
           b_.  Additional sampling stations should be located in areas
               of low hydrologic activity or energy.  The reason for
               sampling these locals is that the lower energy favors the
               settling of smaller sized suspended particulate matter-
               This material, due to the greater surface area per unit
               weight of particulate matter, tends to have higher
               concentrations of associated chemical contaminants.  ' .
               Suggested locations are: (l) on the outside bend of
               channels, (2) in backwater areas or side channels, and
               (3) in areas of heavy shoaling or deposition.
           c_.  Sampling stations should be located in other areas not
               covered in categories a_ and b_ above.  Sampling is neces-
               sary below major point sources and in areas of settling
               to define the maximum concentration that will be found
               in the sediments of the project area.  However, this
               specific property, maximum concentration, would make
               such samples nonrepresentative.  Therefore, samples also
               should be collected at random locations removed or
               upstream from major point sources and in areas of higher
               hydraulic energy (i.e. inside bend of channels).  In this
               way, data obtained from sample analysis will provide
               information on the range of sediment properties and
               compositions that can be expected and the entire set of
               resultant data will be more representative of the project
               area.  The number of sample stations located in such
               areas should be equivalent to the number of stations in
               categories (l) and (2) in b_ above.
           cl.  If a 'control area or a former disposal site is to be
               sampled for comparative purposes, multiple stations
               should be sampled.  Sediment composition from these
               areas will also be variable and cannot be defined based
               on single samples.
           Number of samples.  One of the more difficult tasks is
determining the number of samples that should be collected.  During
                               1-10

-------
the preparation of this manual, persons experienced with the topics of
sediment collection and analysis vere interviewed.   These individuals
could not agree on any specified number of samples.  Responses that
were obtained ranged from "5 replicates at each location" to "more than
50 in a harbor" to "20 samples per hectare."  Again, no specific guidance
can be provided; but several general concepts can be presented.  First,
the greater the number of samples collected, the better the source will
be defined.  Secondly, the mean of a series of replicated measurements
is generally less variable than a series of individual measurements.
Thirdly, statistics generally require two characteristics, usually mean
and standard deviation, because single measurements are inadequate to
describe a sample.  Fourthly, the necessary number of samples is pro-
portional to the source heterogeneity.
           A consideration of the above factors suggests that replicate
samples should be collected at each location and that a minimum of three
replicates is required to calculate standard deviations.  Beyond the
replication at a single point, the factors listed above do not limit
the number of samples needed since it depends on site-specific hetero-
geneity and the desired level of defining the source.  Thus, some other
factor will have to limit the number of samples collected.
           One such factor is financial resources.   In this case, the
number of samples that can be collected and analyzed is determined by
the ratio of available dollars and cost per sample:
              ,,  ,           -,     Dollars for analysis            /.. N
              Numbers of samples =		*	            (1)
                                      Cost per sample
In turn, the cost per sample will depend on the cost of analysis for
each parameter, the specific parameters being analyzed, and the number
of samples to be processed (quantity discounts).  As an aid in estimating
analytical costs, Table 1-1 presents information on cost per analysis,
and Table 1-2 presents information on the number of samples that can be
processed daily.  The average costs in Table 1-1 are based on a 1977
survey of government and commercial laboratories.
           This approach will provide one method of estimating the
number of samples that can.be collected and analyzed.  However, should
the calculated number of samples not be sufficient to establish an

                                 1-11

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                               Table 1-1
                    Analytical Costs by Parameter*

Parameter
Alkyl Benzene Sulfonates
Acidity (total)
Aluminum
Ammonia
Antimony
Arsenic
Bacteria
Barium
Beryllium
Bicarbonate
Bioassay
Biological Oxygen Demand
Boron
Bromide
Cadmium
Calcium
Carbon
Carbonates
Carbon Chloroform Extract
Carbon Dioxide
Chemical Oxygen Demand
Chloride
Chlorinated Hydrocarbons
Chlorine
Chlorophyll
Chr ornate
Chromium

*Personal communication,
Average Cost**
$15.00
k.kQ
10.96
11.81
12.89
16.29
13.97
10. k2
11.25
7.50
Generally
Quoted Upon
Request
27.19
17.31
12.17
9-35
7.56
15.71
7.50
62.50
9.50
Hi. 38
5.79
U2.50
21.05
11.00
8.75
9-05
(Continued)
1977, J. Westhoff, research
Range

$ 2.50
U.OO
3.00
5.00
6.00
2.50
U.OO
U.OO
3.00

10.00
7.00
5.00
2.50
2.00
5.00
3.00
35.00
5.00
7.50
3.00
20.00
1.50
3.00
3.00
IK 00

chemist
of Cost**
_
9.00
- 20.00
- 30.25
- 35.00
- 35.00
- U5.00
- 20.00
- 20.00
- 15-00
^
- 115.00
- 35-00
- 20.00
- 16.50
- 20.00
- 35.00
- 15-00
- 90.00
- 22.00
- 33.00
- 20.00
- 70.00
- 100.00
- 25-00
- 16.50
- 20.00

, WES.
**19TT prices.
(Sheet 1 of 3)
                                1-12

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                         Table 1-1 (Continued)
Parameter
Cobalt
Color
Copper
Cyanide
Detergents
Dissolved Oxygen
Fluorides
Hardness
Hydrogen Sulfide
Hydroxides
Iodine
Iron
Lead
Magnesium
Manganese
Mercury
Methane
Molybdenum
Nickel
Nitrate
Nitrite
Total Kjeldahl
Odor
Oil and Grease
PCB'st
Pesticides
PH
Phenols
Total Phosphorus
Orthopho sphorus
Potassium

Average Cost
$ 9-30
7.30
7.88
19-93
12.50
U.60
12.00
4.85
10.00
7.00
12.00
8.2U
10. U7
8.56
9.20
17.50
15-00
10.88
9.UO
8.17
7.50
18.05
13.67
27.75
33.33
H9.00
3.05
21.11
9.00
5-57
8.77
(Continued)
. Range
$ U.OO
1.50
2.50
5.00
7.50
2.50
3.50
1.50
5.00
3.00
10.00
2.50
2.50
U.OO
2.50
11.00
11.00
U.OO
2.50
3.00
5.00
6.00
1.00
7.00
25.00
20.00
1.50
5.00
6.00
2.00
2.00

of Cost
- 9.30
- 25.00
- 20.00
- Uo.oo
- 20.00
- 8.00
- 25.00
- 10.00
- 15.00
- 15.00
- 15.00
- 20.00
- 20.00
- 20.00
- 20.00
- 35.00
- 35.00
- 20.00
- 20.00
- 16.50
- 12.00
- 30.25
- 30.00
- 150.00
- Uo.oo
- 75-00
5.00
- 36.50
- 12.50
- 10.00
- 20.00

t Polychlorinated biphenyls.
(Sheet 2 of 3)
                                1-13

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Table 1-1 (Concluded)
Parameter
Selenium
Silica
Silver
Sludge Volume Index
Sodium
Total Solids
Volatile Solids
Dissolved Solids
Suspended Solids
Settleable Solids
Specific Conductance
Specific Gravity
Strontium
Sulfate
Sulfide
Sulfite
Surfactants
Tannin and Lignin
Taste
Thallium
Thiocyanate
Tin
Turbidity
Vanadium
Volatile Acids
Zirconium
Zinc
Average Cost
$20.72
10.03
10.70
12.00
8.95
7.00
6.63
7-23
7-00
5.85
3-95
8.13
20.79
8.75
11.23
7.89
18. OU
1U.86
50.00
12.20
12.50
11.15
3.89
11.11
20.00
21.67
9-63
Range
$ 5-00
it. 00
U.OO
10.00
2.00
U.50
1.50
1*. 50
2.00
2.00
1.50
1.50
U.oo
U.OO
5.00
3.00
10.00
7.50
20.00
U.OO
10.00
2.50
1.50
U.OO

10.00
2.50
of Cost
- 55.00
- 20.00
- 20.00
- lU.OO
- 20.00
- 11 . 00
- 16.50
- lU.OO
- 13.50
- 12.00
5-00
- 20.00
- 75.00
- 18.00
- 25.00
- 12.50
- 28.00
- 25.00
- 100.00
- 20.00
- 15.00
- 20.00
6.00
- 30 . 00
-
- 35.00
- 20.00
                                (Sheet  3 of  3)




       1-lU

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                               Table 1-2
                      Performance Requirements,*
                   Number of Samples Per Day Per Man
                                                       Number of Water
	Parameter	                     Samples per Day
Dissolved Oxygen (probe)                                  150
Dissolved Oxygen (Winkler)                                120
pH                                                        175
Conductivity                                              120
Turbidity (Jackson)                                        75
True color (filtration)                                    60
Oxygen uptake
Biological Oxygen Demand (probe)                           ho
Biological Oxygen Demand (Winkler)                         30
Immediate Oxygen Demand
Chemical Oxygen Demand                                     2k
Chlorine Demand
Alkalinity (total)                                        100
Acidity (total)                                           100
Total Kjeldahl Nitrogen (manual)                           20
Total Kjeldahl Nitrogen (automated)                       100
Phosphorus - Total (manual)                                50
Phosphorus - Total (automated)                            100
Phosphorus - Total Soluble                                100 - 150
Solids - suspended and dissolved                           20
Solids - total and volatile
Phenol (distillation)                                      20
Oil and Grease (Soxhlet)                                   12
NH3-N (automated)                                         100
N03-N (automated)                                         100
N02-N (automated)                                         100
Chloride (automated)                                      100
                              (Continued)
* Personal communication, 1977, J. Westhoff, research chemist, WES.
                                  1-15

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                         Table 1-2 (Concluded)
                                                        Number of Water
	Parameter	         Samples per Day
Sulfate (automated)                                        100
Magnesium (direct aspiration)                              100 - 150
Silica (automated)                                         100
Sodium (direct aspiration)                                 150
Potassium (direct aspiration)                              150
Calcium (filtration/direct aspiration)                     100
Arsenic                                                     20
Fluoride (distillation)                                     25
Fluoride (automated)                                       100 - 125
Cyanide (distillation)                                      12
Sulfide (titrimetric)                                       50 - 75
Manganese (direct aspiration)                              150
Total Iron (digestion/direct aspiration)                    60 - 80
Copper (direct aspiration)                                 150
Cadmium (direct aspiration)                                150
Nickel (sample concentration/direct aspiration)            150
Zinc (direct aspiration)                               '    150
Lead (direct aspiration)                                   150
Chromium (direct aspiration)                               150
Pesticides
   Chlorinated                                          10 per 12 days
   Sulfated                                             10 per 12 days
   Carbon Filter                                        10 per 18 days
                               1-16

-------
adequate sampling program (i.e. number of samples not sufficient to

allow triplicate sampling at all locations indicated in the section on

Sampling Site Locations), one of the following trade-offs will have to

be accepted:*

           a_.  Reduce the replicate sampling at each station.  This
               will allow the chemical distribution within the project
               area to be determined; but variability at a single
               sampling location cannot be calculated.

           b_.  Maintain replicate sampling but reduce the number of
               sampling locations.  This will result in the project
               area being less well defined but sampling variability
               can be calculated.
           c_.  Reduce the number of analyses that will be run on each
               sample.  In this way, samples only have to be analyzed
               for specific parameters of concern in a given project
               area.  Because the analyses to be run are site specific,
               no mandatory list of analyses can be recommended at
               this time.

           d^.  Increase the financial resources available for sample
               analysis.  This will increase the number of samples
               that can be collected and analyzed.

           A second factor that can be used to estimate the number of

samples needed for a project is the level of statistical reliability
or confidence that is desired in the results.  In the case where a

random sample has been taken and in reference to any particular
chemical parameter of interest, this can be calculated as:  * 1

                              n  = t2 s2                           (2)
                                   -
where
     n   =  number of samples

     t2  =  student's-t distribution value

     s   =  population variance
      n
     d   =  statement of margin or error
* The distinction between option a_ and option b_ should be based on
  project-specific goals.  If option &_ is used (more stations,
  fewer replicates), the results will provide a better indication
  of distribution patterns in the project area (Synoptic Survey);
  but it will be difficult to compare individual stations.  On the
  other hand, if option b_ is used (fewer stations, more replicates),
  the results will provide -a better indication of variability at
  one location and a comparison between sampling stations.  However,
  the project area will be less well defined.

                                 1-17

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It should be noted that this method will most likely lead to different
numbers of samples being required for each parameter and trade-offs  will
have to be made in selecting the final number of samples to collect.
           Because of the inherent heterogeneous nature of sediments,
one should be prepared to accept rather large values for  n   or  d
when using this approach.  As an example, in one study in Lake Erie,12
100 samples were collected from a 100-square-mile area.*  Based on  n   =
                                                                    o
100, t2 = k (95 percent confidence level), and s2 = 1*6.10 (observed
variance = 6.79)5 the calculated  d  value is l.h mg/kg.  In other
words, based on the analyses of 100 samples, one could be 95 percent
confident that the mean cadmium concentration was within l.k mg/kg of
the true mean.  If one wanted to be 95 percent confident that the mean
cadmium concentration was within 1.0 mg/kg of the true mean, a total
of 18^ samples would have to be collected.
           This approach obviously requires a knowledge of historical
data and/or an examination of the project site.  It can be used to
calculate the required number of samples in a project area after
stating the confidence desired in the final data  d  and knowing the
                                  n
variability in the project area  s  .  However, the approach can also
be used after completion of a sampling program, when  n   and  s2 are
known, to calculate the level of confidence that can be placed in the
final data.
           It is suggested that some consideration be given to
collecting samples (locations and numbers) in excess of that determined
by either of the above processes.  The samples do not have to be
scheduled for analysis and may even be discarded later without analysis.
However, sediments can be highly heterogeneous as discussed earlier;
and should sample analysis indicate some sort of anomalous results,  it
is easier to analyze additional samples already on hand rather than  to
remobilize a field crew.  Also, the additional variable of different
sampling times is avoided with this approach.
           Frequency of sampling.  Frequency of sampling will depend
on the available resources and the size of the project.  Most persons

* A table of factors for converting U. S. customary units of measure-
  ment to metric (Si) is presented on page xv.

                                1-18

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interviewed during the preparation of this manual suggested that
seasonal fluctuations of sediment concentrations may not be critical
and a single sampling prior to a dredging or filling operation may be
sufficient for a new work project.  A sampling frequency of once per
year would probably also be sufficient for an annual maintenance
project, unless there is a "reason to believe" otherwise (i.e., some
major change in point sources or basin hydrology).
           An assessment of sampling frequency will be subjective.
The factors that are likely to influence the adequacy of the sampling
frequency are duration of the project, time of the year, and local
activities.  The shorter the project duration, the less likely that
multiple samplings would be required.  However, as the project
duration increases, particularly to time frames of three months or
longer, consideration should be given to increased sampling frequency.
The second two points are an extension of the first factor.  There may
be no potential problems when a project is initiated; but a lengthy
project may extend into a critical spawning or migration season.  This
could force an increased sampling frequency or, at a minimum, a
reevaluation of the earlier data based on changes in seasonal activities.
Obviously, considerations of this type are site and project specific.
It is recommended that an increased sampling frequency be used when
a project is expected to last longer than one quarter (three months),
unless it is known that the project will not impact or be impacted by
a locally important seasonal activity.
Sampling techniques selection
           Sampling equipment is frequently selected based on reli-
ability, efficiency, and contamination potential.  Information on these
characteristics is summarized in Section 2.  Project managers should
also be aware of the fact that the selection of sampling equipment may
affect the apparent sample variability.  As discussed earlier, sample
variability can influence the number of samples required for analysis
or the confidence that can be placed in the resultant data.
           Sediments are frequently stratified in the vertical dimension
as well as in the horizontal dimension.  This source of variability
                                 1-19

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should be considered when establishing a sampling program, particularly
when chosing a method of sampling (i.e., grab or dredge vs. corer).   A
grab sampler (i.e. Ponar, Ekman, and Orange Peel) is a device that usually
triggers after free-falling and is used to retrieve surficial sediments.
The difficulty with this approach is that the depth of sediments pene-
trated by the sampler may vary, depending on the weight and shape of the
sampler, the sediment texture and density, the height of free-fall, and
the angle of impact.13'*1*  Since many chemical constituents display
surface enrichment trends, the use of grab samplers may introduce
analytical variability into the final data that is a function of dredge
pentration rather than being a property of the sample.
           For example, in a hypothetical situation in which the
concentration of a constituent varies from 1 to 10 mg/kg, an average
concentration as a function of dredge penetration was calculated
(Figure l-l). Al-cm surface grab in this example would have an
average concentration of 10, while a 3-cm grab would have an average
concentration of 5-6; and a sample that penetrates more than 8 cm
would have an average concentration of less than 3.  Thus, in an
extreme case, a differential dredge penetration could produce more
than a 300 percent variation in the analytical results at the same
sampling location.  A difference in penetration of 1 cm could produce
analytical variability of 7 to k3 percent.  The actual variation will
depend on the site-specific depth profile and the differential depth
of penetration.
           A study by Skoch and Britt15 demonstrated the effect of
corer vs. dredge sampling.  Dredge samples collected over a 2-year
period had an average phosphate concentration of 2,2k +_ O.kk mg/kg.
The uppermost section of a core (0 to 2.5 cm) sample had an average
phosphate concentration of 2.73 ;t 0..29 mg/kg; and a lower section of
the core (12.5 to 15 cm) had an average phosphate concentration of
2.17 +. 0.2k mg/kg.  Thus, the analytical results obtained with an
Ekman dredge were not directly comparable to the corer results; and,
in addition, the dredge results were more variable as suggested by the
larger standard deviation.  It was concluded that core samplers may be
more valuable than dredges for sediment studies.

                                1-20

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                        RELATIVE CONCENTRATION, ing/kg


                         k          6          8         10
12
   2.
o
w
Q
   6.
   8-
  10-
Depth of
Sample
Collected
cm
1
2
3
k
5
6
7
8
9
10
Calculated
Average
Concentration
mg/kg
10
7
5.6
4.7
4.0
3.5
3.1
2.9
2.7
2.5
          Figure  1-1.    Average concentration as a function of

                         dredge penetration
                                  1-21

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           The previous discussion assumes that the vertical strati-
fication in sediments is uniform.   However, irregular folding of sediment
layers frequently occurs.  This situation can make it more difficult
to compare grab samples taken from the same area and compound the
potential analytical "bias identified earlier.
           The choice between grab samplers and corers can be based on
convenience or availability when the deposit to be sampled is homogeneous
with depth since both methods should give the same result.  However,  the
selection of a grab sampler in a stratified deposit is liable to yield
biased conclusions and is certain to lose potentially valuable infor-
mation.  It also is cautioned that such use may require the collection
and analysis of more samples to compensate for the analytical bias that
may be introduced.  Thus, it is suggested that corers would be the
preferential method of sample collection and should be used where
available.
           One situation where the selection between grabs and corers
may not be critical is in the evaluation of dredging activities in
maintenance work projects.  In these areas, the sediments that have
accumulated since the last maintenance project are generally subjected
to continual reworking due to marine traffic.  The net effect of this
activity homogenizes the sediments that have accumulated.  Because
maintenance dredging is concerned with the removal of accumulated
sediments rather than deepening or creating new channels and the
previous discussion indicated no difference between corers and dredges
in homogeneous deposits, grab samplers should be sufficient in these
situations.
           When the project being evaluated includes either deepening
of an older channel or creation of a new channel, it is recommended
that cores be collected.  Also, when possible, the cores should be
taken to a depth equivalent to the proposed project depth.
Sample preservation
           Methods.  The importance of sample preservation between time
of collection and time of analysis cannot be overemphasized.  The
purpose of collecting samples is to gain understanding of the source  of
                               1-22

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the samples; any changes in sample composition can invalidate conclusions
regarding the source of the samples.1*  To phrase it another way,  results
based on deteriorated samples negate all efforts and costs expended to
obtain good samples.
           The most efficient way to ensure a lack of sample deteriora-
tion is to analyze samples immediately.  However, this is usually not
pratical because of:
           a_.  The number of samples collected.
           b_.  The type of equipment needed for analysis.
           c_.  The steps involved in sample preparation.
Therefore, some method must be relied upon to extend the  integrity of
the sample until the analyses can be completed.  In taking this
approach, it must be remembered that complete stabilization is not
possible and no single preservation technique is applicable to all
parameters.
           Preservation methods are relatively limited and are generally
intended to retard biological action, hydrolysis, and/or  oxidation
of chemical constituents and reduce volatility of constituents.15  The
methods are limited to pH control, chemical addition, sample isolation,
and temperature control (refrigeration and/or freezing).   Selection of
a preservative should be based on the purpose of the study and the
constituent to be measured.  If one is interested in the  total con-
centration of iron in sediments, either drying, freezing, or refrigera-
tion in an airtight container would be satisfactory.  However, if one
is concerned about the mobilization of iron from sediments to the
water column, only the latter preservation technique, refrigeration
in an airtight container, would be acceptable.  Because of the limited
number of preservation techniques, it is obvious that they are not
parameter specific.  The use of one preservative may invalidate the
use of that sample for the analysis of a second parameter (i.e.,
samples preserved with mercury salts to retard biological activity
cannot be analyzed for mercury; samples preserved by drying should
not be analyzed for oxygen demand).  As a consequence, multiple
samples will have to be collected and individually preserved or a
single sample will have to be split into subsamples and preserved as

                                 1-23

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required.  The elapsed time between sample collection and sample
preservation must be kept to an absolute minimum.
           Appropriate containers and methods of preservation for water
samples have been generally agreed upon and are presented in Sections 2
and 3 of this handbook.  Transportation and preservation of sediment
samples have not been fully evaluated.  However, it is suggested that
glass containers be used when the sample is to be analyzed for organic
constituents and glass or plastic containers be used when the sample is
to be analyzed for inorganic constituents as suggested for water
samples.16  Sediment samples should be sealed in airtight containers to
preserve the anaerobic integrity of the sample and maintain the solid
phase-liquid phase equilibrium.
           Sample handling.  Proper sample handling is essential to
obtain successful results from any monitoring program.  It is the
responsibility of the project manager to ensure that samples be
correctly handled between collection and analysis.  This includes:16
           a_.  Using noncontaminating sampling devices.
           b_.  Having appropriately cleaned sample containers
               available (glass for organic analysis, acid-rinsed
               bottles for metals, etc.).
           c_.  Having appropriate chemical preservation and/or
               preservation techniques readily available.
           d_.  Using a reliable sample labeling-and-identification
               procedure.
It is also necessary that the samples be logged in after collection and
analyzed within prescribed time limits.  Each of these factors is
specified or discussed in more detail in Section 2 but are mentioned
here because of their importance.
                           Quality Control

           The factors associated with sample handling are important
because they influence the quality of the data being generated.  It
should be apparent, based on the factors mentioned above, that an
effective quality control program must be an integral part of a
project from the initiation of field sampling and should be considered

-------
during project planning.  During the initial meeting,  the field crew
should be made aware of the fact that chemical changes can occur
following collection of samples; they should also know how to handle
the samples to minimize or prevent these changes.  In  addition, it
may be helpful if the field crew knows the type of analyses to be
performed so they can minimize sample contamination problems.  At the
same time, laboratory personnel should be reminded of  their responsi-
bility to complete the required analyses within the specified time
period.
           A complete quality control program should emphasd ze two
areas.  The first area is sample handling techniques.   This is
necessary because the greatest potential for sample deterioration
and/or contamination occurs during the preanalysis steps of sample
collection, handling, preservation, and storage.  These problems can
be minimized by following prescribed sample handling techniques.
The second area to be emphasized is that of analytical quality control.
This is accomplished by analyzing field replicates, split samples, and
spiked samples.    For water samples, the quality control program
should include the analysis of field replicates, samples spiked in
the field, laboratory duplicates, and samples spiked in the laboratory.
For sediment samples, the quality control program should include the
analyses of field replicates, laboratory duplicates, and samples
spiked in the laboratory.  Field-spiked sediment samples would be
awkward since the spike would probably not equilibrate and mixing
would destroy sample integrity.  A recommended quality control program
should consist of 15 to 20 percent of the total sample load.   It is
further recommended that the laboratory participate in quality control
round-robin sample analysis studies.

                       Additional Considerations

           The purpose of conducting a sampling effort is to gain
information.  The quality of the information gained can only be as
good as the care in preparing for and implementing the effort.  Since
                                1-25

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definitive guidance on the number of sampling locations and number of
samples cannot be given, one step to ensure a high quality field program
is to use experienced field personnel.   In addition to experience,
another useful characteristic is a high level of communication.   Field
personnel should be familiar with the purpose of the study and the
analyses that will be completed on the samples.   This will permit them
to use their judgment in the most constructive manner and alert  them
to specific precautions that should be taken in sample collection,
preservation, and storage.

                                Summary

           This section has endeavored to present the major factors to
be considered in implementing a field sampling program.  Factors such
as sample handling and quality control should be part of standard
laboratory practice and are mentioned for completeness.  Other factors
such as method of sampling (core vs. dredge) and type of test (standard
elutriate test vs. elemental partitioning vs. bulk analysis) are
mentioned because of the choice afforded.  Each alternative provides
certain information and sacrifices other information.  Specific
guidance can be provided for each procedure; but it is up to the
project manager to decide which procedure is most useful for his
project.  Finally, there are factors such as sampling locations  and
number of samples for which specific guidance cannot be given because
of site-specific influences.  Again, the project manager will have to
address these factors; the potential trade-offs  that are involved have
been presented herein for consideration (greater replication vs.
fewer sampling locations; more samples and fewer analyses per sample
vs. greater sample characterization of fewer samples).
                                 1-26

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                              References
 1.  Environmental Protection Agency/Corps of Engineers Technical
     Committee on Criteria for Dredged and Fill Material.   "Ecological
     Evaluation of Proposed Discharge of Dredged Material  into Ocean
     Waters; Implementation Manual for Section 103 of Public Law
     92-532 (Marine Protection, Research, and Sanctuaries  Act of 1972)."
     Environmental Effects Laboratory, U.S. Army Engineer  Waterways
     Experiment Station, Vicksburg, Mississippi.  2k p. +  APP.  July
     1977 (2nd Printing 1978).
 2.  Griffiths, J. C.   Scientific Method in Analysis of Sediments.
     McGraw-Hill Book Company, New York.  508 p. (1967).
 3.  American Public Health Association.  Standard Methods for the
     Examination of Water and Waste Water Including Bottom Sediments
     and Sludges.  APHA, New York.  769 p. (1965).
 U.  Research and Education Association.  Modern Pollution Control
     Technology. Vol.  2. Water Pollution Control and Solid Waste
     Disposal.  Research and Education Association, New York.
     Unnumbered (1978).

 5.  Griffiths, J. C.   "Statistical Methods in Sedimentary Petrography."
     In:  Milner, H. B. (Ed.), Sedimentary Petrography. George Allen
     and Unwin, Ltd.,  London,  pp. 565-617 (1962).
 6.  Sherma, J.  "Manual of Analytical Quality Control for Pesticides
     and Related Compounds in Human and Environmental Samples."
     Contract 68-02-1727-  U. S. EPA Health Effects Research Laboratory,
     Research Triangle Park, North Carolina.  EPA-600/1-76-017.
     Unnumbered (1976).
 7.  Helmke, P. A., Koons, R. D., Schomberg, P. J., and Iskandar, I. K.
     "Determination of Trace Element Contamination of Sediments by
     Multielement Analysis of Clay-Size Fraction."  Environmental
     Science and Technology 11: 98^-989 (1977).
 8.  Forstner, U., Patchineelam, S. R., and Deurer, R.  "Grain Size
     Distribution and Chemical Associations of Heavy Metals in Fresh-
     water Sediments (Examples from Bodensee and Rhine).    Presented
     before ACS Environmental Chemistry Division, 175th National
     Meeting, Anaheim, California.  Unnumbered (1978).
 9.  Federal Register. Vol. ^0, No. 173.  Friday, 5 September 1975-
10.  Sydor, M.  "Particulate Transport in Duluth-Superior  Harbor."
     Physics Department, University of Minnesota, Duluth,  Minnesota.
     Grant R-803592.  Draft Report.  101 p. (1978).
11.  Weber, C. E. (Ed.).  "Biological Field and Laboratory Methods  for
     Measuring the Quality of Surface Water and Effluents."  National
     Environmental Research Center, U. S. EPA, Cincinnati, Ohio.
     EPA-670A-73-001.  Unnumbered (1973).
                                1-27

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12.  Great Lakes Laboratory.  "Airport Feasibility Study for the Lake
     Erie Regional Transportation Authority."  Contract Report prepared
     for Howard, Needles, Tammen, and Bergendoff.  Report 6-5.  Great
     Lakes Laboratory, State University College at Buffalo, Buffalo,
     Nev York.  278 p. (197*0 .
13.  Hudson, P. L.  "Quantitative Sampling with Three Benthos Dredges."
     Trans. Amer. Fish Soc.  99:  603-607 (1970).

1^.  Christie, N. D.  "Relationship Between Sediment Texture, Species
     Richness, and Volume of Sediment Sampled by a Grab."  Marine
     Biology 30: 89-96 (1975).
15.  Skoch, E. J. and Britt, N.  W.  "Monthly Variation in Phosphate
     and Related Chemicals Found in the Sediment in the Island Area of
     Lake Erie, 1967-1968 with Reference to Samples Collected in 1.96k,
     1965, 1966."  Proc.  12th Conf. Great Lakes Res.  pp. 325-3^0
     (1969).
16.  U. S. Environmental Protection Agency.  "Methods for Chemical
     Analysis of Water and Wastes."  National Environmental Research
     Center, U. S. EPA, Cincinnati, Ohio.  EPA-625/6-7^-003.  298 p.
     (197M.
                                1-28

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                 SECTION 2:  FIELD/LABORATORY GUIDANCE

           This section is intended primarily for the field and labora-
tory personnel that will be implementing the sampling program.   The
nature of the guidance in this section is less subjective than the
previous section because it deals with how to perform a test rather
than when or on what samples to perform a test.   The subjects  covered
include sample collection, sample container preparation, sample
handling and storage, quality control, and sample testing procedures.
Each of these subjects require some selection of options (grab sample
vs. core sample; freezing, drying, or moist storage of samples; use
of bulk analysis, elutriate testing, and/or elemental partitioning).
However, the choice of the various options should be addressed in the
project planning meeting as discussed in Section 1.  The purpose of
this section is to provide guidance on how to handle the samples or
perform a specific test after it has been decided to use a specific
procedure.
           This section follows the logical progression of sample
collection, sample handling and storage, and testing.  This represents
the physical sequence of handling samples.  However, it should be
obvious that preparations for sample storage and analysis must be
made prior to sample collection in order to minimize the undesirable
effects of sample contamination and alteration between sample collection
and sample analysis.

                      Method of Sample Collection

           The type of samples to be collected will depend on the
specific testing procedures to be used which, in turn, should be
based on the purpose of the project since each procedure provides
discrete information on the sample.  Water samples will furnish
information on existing water quality at a proposed project site.
Water and sediment samples are required to perform an elutriate test
in order to estimate chemical mobility during dredging and disposal
activities.  Sediment samples are required to determine the chemical

                                  2-1

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form distribution (elemental partitioning or fractionation)  or the
total concentration (bulk analysis) of a particular constituent at the
collection site.
Water samplers
           Devices available for the collection of vater samples can be
classified in two categories:  discrete samplers and pumps.   While
either device can be used satisfactorily, it should be remembered
that sampling equipment should be constructed of noneontaminating
material.
           For collecting vater samples, free flushing, messenger-
triggered stainless steel bottles with no gaskets, and teflon-coated
bottles with teflon seals are the better choices from among  those
available.1  Polyvinyl chloride (PVC) water bottles can also be used
provided the 0-ring gaskets, if rubber, are replaced with teflon or
some other noncontaminating material.
           Water samplers are subject to contamination as the open
bottle passes through the water's surface where organics, other
floatables, and metals have been shown to accumulate.1  This potential
problem can be minimized by avoiding areas with visible surface slicks
or films.  In addition, the contaminating effects are further reduced
by the natural flushing action of the sampler and the selection of a
sampler with a large volume-to-surface area ratio.  If surface films
are a recurrent problem, an alternative collection method is to use a
sampler that is closed during descent and only opened at the desired
depth.
           When pumps are selected for water sample collection, care
should be taken to run the apparatus for a sufficient length of time
to ensure that the system has been thoroughly flushed in order that
the collections are representative of the sampling location.  Allowing
a water volume equal to three times the combined tube volume to pass
through the system before collections are retained should be sufficient
for this purpose.  A point of concern should be the contamination
potential of packing and lubricants used with the pump.  These effects
can be avoided by using peristaltic pumps, magnetically coupled
impeller designs, or vacuum pump collectors.  However, peristaltic

                                   2-2

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pumps and magnetic pumps should "be considered superior to the use of
vacuum pumps as the latter will degas the sample.   This could, in turn,
cause other secondary chemical changes in the sample being collected.
Sediment samplers
           There are three broad classifications of sediment collecting
devices.  These are corers, grabs, and dredges.  Corers generally
produce the least disturbed samples; grabs collect larger surface
samples; and dredges collect larger, well-mixed samples that are
considered qualitative.1  Because there are concerns about the repre-
sentativeness of dredge samples, dredges should only be used where
other sediment samplers are not applicable.
           The choice between corers and grabs should be based on the
type of project being evaluated.  If the project requires new work
or harbor deepening, sediment samples should be collected with a
corer.  In this way, samples can be collected of all the material
to be involved in the proposed project.  If the project is classified
as maintenance work, grabs will usually be sufficient.  Practical
experience has shown that hydrologic conditions at such locations
usually result in homogeneous sediments due to currents and/or marine
traffic.*  Since vertical stratification would not be expected and
older sediments would not be disturbed, coring devices generally are
not required for sample collection under these conditions.
           Corers function by driving a tube into the sediments,
usually through the use of gravity, hydrostatic pressure, or
vibration.1'5  The length of core that can be collected generally is
limited to 10 core diameters in sand substrate and 20 core diameters
in clay substrate.  Longer cores can be obtained but substantial
sample disturbance results from internal friction between the sample
and the core liner.
           Free-fall cores can cause compaction of the vertical
structure of sediment samples.  Therefore, if the vertical strati-
fication in a core sample is of importance or interest, a piston
* Personal communication, 1978, from R. Bowden, Chief, Great Lakes
  Surveillance Branch, Great Lakes National Program Office, EPA
  Region V, Chicago, Illinois.
                                 2-3

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corer should be used.  These devices utilize both gravity and hydro-
static pressure.  As the cutting edge penetrates the sediments, an
internal piston remains at the level of the sediment-water interface,
preventing sediment compression and overcoming internal friction.  If
samples will not be sectioned prior to analysis, compaction is an
academic problem; and free-fall corers are a suitable alternative.
           Grabs are designed primarily to retrieve surficial
sediments.  A problem identified with the use of grab samplers is that
shock waves are generated ahead of the descending samplers and these
waves can wash away light, unconsolidated sediments and unattached
benthic organisms.1'5 8  This effect is minimized by using samplers
with flaps, screens, or valves to create a flow-through system during
descent.  An added benefit is that sample washout protection is
provided by the flaps during recovery.1'5
           Another problem with grabs is that sampling characteristics
(sediment penetration) can be influenced by bottom hardness, depth
to sample, and fall rates and angles as well as lateral vessel
motion.9  These factors can contribute to apparent sample composition
variability where sediment concentrations vary vertically.  Therefore,
it is recommended that the use of grab samplers be restricted to areas
of vertically homogeneous sediments.
           The various sediment samplers were evaluated for sampling
efficiency, reproducibility, and sample protection.5  The observations
and evaluations are presented in Tables 2-1 and 2-2.  Based on these
studies, the Shipek and Ponar dredges would be considered the best
sampling devices for maintenance projects.

                           Sample Collection

           Sample site locations and number of samples to collect are
project specific.  The factors that influence the decision on where
to sample and how many samples to collect are project purpose(s),
major point sources in the project area, activities in the project
area, hydrologic conditions, and sample variability.  The project
                                2-U

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                               Table 2-1
               Operational Evaluation of Grab Samplers*
       Grab
Franklin Anderson
Dietz-LaFond
Birge-Ekman
           Trigger System Reliability
Petersen

Ponar

Shipek
Franklin-Anderson
Dietz-LaFond

Birge-Ekman


Petersen



* After Sly.'
Good, but perhaps too sensitive on hard sand and
 gravel bottom.

Poor, unless area of trigger foot is increased to
 at least 50 cm2.  Triggering may often be
 impossible in very soft mud unless the foot has
 been modified.

Good.  Triggered by messenger weight dropped
 from surface, normally consistent but can be
 affected on soft bottoms if sampler is allowed
 to settle for too long before dropping the
 messenger.
Fair to good, though tends to be a little over-
 sensitive on hard sand and gravel bottoms.

Good, though like the Petersen, tends to be a
 little oversensitive on gravel bottoms.
Good, though some slight settlement may occur
 before triggering on very soft materials.
 Sampler may fail to trigger when lowered
 gently on soft bottoms.  By lifting and dropping
 the trigger weight a few centimeters after
 bottom contact, abortive casts may be avoided.
 The slight movement of the inertial trigger
 weight has no other effect on the sampler.

	Jaw Shape, Design, and Cut	

Poor.  During the first stages of closure and when
 under the greatest pressure of springs and weight,
 the jaw shape loosely follows the arc of cut.
 However, the degree of fit becomes progressively
 worse as the closing pressure is reduced.  Because
 each jaw is semicylindrical in shape, sample
 displacement is necessary within it if anything
 near maximum capacity is to be achieved.
Poor.  As for Franklin-Anderson.

Excellent.  Jaw shape exactly follows arc of cut
 and almost no sample displacement occurs.

Poor.  Comments as for Franklin-Anderson, except
 that, instead of the reduction in closure pressure
         (Continued)

                                     (Sheet 1 of it)
                                   2-5

-------
                         Table 2-1 (Continued)
        Grab
Petersen (Continued)



Ponar

Shipek
          Jaw Shape, Design,  and Cut
Franklin-Anderson
Dietz-LaFond
Birge-Ekman
Petersen
Ponar
Shipek
 being produced by slackening of tensional
 springs, the same result is effected by
 reduced leverage on the scissor arms mounted
 across the hinge line.
Excellent.  Jaw shape exactly follows arc of
 cut and almost no sample displacement occurs.
Excellent.  As for Ponar.  In addition, the
 rotation of the bucket is extremely rapid.
 In most cases the rotational shear is far
 greater than the sediment shear strength,
 thus the cutting action is very clean
 (producing minimal disturbance), particularly
 in soft clays, muds, silts, and sands.

  Preservation and Protection from Washout

Fair, but the tightness of closure is largely
 dependent upon the lack of grains trapped
 between the edges of the jaws.  Providing a
 tight fit between the two jaws is obtained,
 the sample is well shielded against washout.
 If the jaws are kept open by material
 trapped between the jaws, washout can be
 severe or total.
Fair.  Comments as for Franklin-Anderson.
Good, except when the sampler is used in very
 coarse or shelly sediment.  Under these
 conditions, material may be trapped between
 the jaws, preventing their closure.  In this
 case, washout may be severe.  The jaws are
 so designed that they slightly overlap one
 another, thus a slight imperfection of
 closure can be tolerated.
Good.  Comments as for Birge-Ekman.
Good.  Comments as for Birge-Ekman.  In
 addition to the overlap jaws, this sampler
 has a pair of metal side plates, mounted
 close to the moving side faces of the jaws.
 These plates further reduce the possibility
 of washout.
Excellent.  The great advantage of the Shipek,
 over all of the other samplers described, is
                              (Continued)
                                (Sheet 2 of k)
                                  2-6

-------
                         Table 2-1 (Continued)
       Grab
Shipek (Continued)
Franklin-Anderson
Dietz-LaFond
Birge-FJonan
Petersen
    Preservation and Protection from Washout	

 that the bucket closes with its separation
 plane aligned in the horizontal rather than in
 the vertical.  Good samples can be retrieved
 even when bucket closure is prevented by
 pebbles or similar material, even 2 to 5 cm
 across.  With the bucket properly rotated,
 washout is completely avoided.

	Stability	

Fair.  Despite the weight of this grab, it tends
 to "stream" at an inclined angle under conditions
 of rapid ship drift or fast water flow.
 Provided lowering conditions are calm and
 stable, the sampler will hold upright during
 the initial sampling process; if, however, the
 line is allowed to slack, the sampler will fall
 over.
Poor.  This sampler is very sensitive to "strea-
 ming" and will rarely operate in the vertical
 position unless used in ideal conditions.
 Its tendency to maintain an inclined attitude
 during descent sometimes results in a failure
 to trigger.

Fair-.  Despite the light weight of this sampler
 and its tendency to "stream," its wide base
 gives good stability and stance once it has
 come to rest on the sediment floor.  Under poor
 sampling conditions, however, it becomes
 impossible to operate because:  (a) the
 sampler, due to its light weight, is continually
 being lifted and dropped and "streamed" along
 the bottom, and (b) any slack in the line,
 particularly near the sampler, is likely to
 impede the proper function of the triggers'
 messenger weight.  It tends to roll over after
 triggering on all but soft bottoms.
Good.  This is a heavy sampler with a wide base
 line (when the jaws are open).  It maintains a
 near vertical descent under all conditions,
 but after sampling it tends to fall over
 (unless on a soft bottom).
                              (Continued)

                                   2-7
                                                           (Sheet  3 of h)

-------
                         Table 2-1 (Concluded)
       Grab	    	Stability	

Ponar                   Very good.  Comments as for Petersen; because
                         of its weight and wide baseline (when jaws
                         are open), this grab has a good vertical
                         descent under most conditions and has a
                         stable stance on the bottom.   The presence of
                         the fixed side plates prevents the grab from
                         falling over after jaw closure and helps in
                         preserving a near perfect bottom sample.

Shipek                  Excellent.  Despite the large size of this
                         sampler, its weight ensures a near perfect-
                         vertical descent even under conditions of
                         rapid drift or fast water flow.  The sampler
                         is also very stable even on bottom slopes 20
                         degrees or more.  This stability ensures the
                         minimum possible disturbance of the sample
                         material.
                                                        (Sheet  h of k)
                                 2-8

-------
                               Table 2-2
         Operational Suitability of Corers and Grab Samplers*
      Sampler
Benthos Gravity
 Corer
Alpine Gravity
 Corer
Phleger Corer
Multiple Corers
Franklin-Anderson
 Grab
Dietz-LaFond Grab
Birge-Ekman Dredge
                 Characteristics
Cores of 3 m-or less in soft clays, muds, or
 sandy silts.   Particularly suitable for studies
 of the sediment/water interface, for studies on
 depositonal sediment structures.

Cores of 2 m or less in almost all sediment types.
 The rugged nature of this corer lends itself to
 general usage.  For studies involving sediment
 structure or large volumes of material, the
 corer is unsuitable; for studies of a pilot
 nature, or to prove the suitability of an area
 for piston coring, this gravity corer is excellent.
Cores of 0.5 m or less, in almost all sediment
 types.  Particularly suited to bottom materials
 containing a high percentage of fiberous organic
 material.  The low cutter angle, the narrow wall
 thickness and high point loading, and the
 extremely sharp cutter, make it very suitable
 for sampling shallow lacustrine and estuarine
 deposits, marsh deposits, and thin peat beds.

Still under investigation.
Suitable for obtaining material for bulk sample
 analysis.  Works best in soft clays, muds, silts,
 and sands.  Will occasionally obtain a good
 gravel sample.  Material of no use for structural
 or other specific analyses.

Can be used for general sampling but not
 recommended for any particular use.  Of all the
 samplers tested, this pattern proved to be the
 least suitable.

Suitable for soft clays, muds, silts, and silty
 sands.  This sampler should be used under calm
 water conditions, typically in small lakes or
 restricted areas.  The lack of sample disturbance,
 square cross section, and moderate penetration
 make this sampler suitable for detailed studies
 (i.e. biological and geochemical) of the top
 2 to 3 cm of bottom sediment.  Because of its
 light weight and easy handling, it is well
 suited to small boat operations.

        (Continued)
* After Sly.
                                  2-9

-------
                         Table 2-2 (Concluded)
   Sampler
                   Characteristics
Petersen Grab
Ponar Grab
Shipek Bucket
 Sampler
This sampler, like the Franklin-Anderson, is suitable
 for taking bulk sample material in most types of
 sediment.  It is quite unsuited for studies of
 detailed and specific sediment properties, though
 it is perhaps a little more successful in taking
 gravel samples.  Either of these tvo samplers
 (Petersen or Franklin-Andersen) will do well as a
 general purpose bulk sampler.
An excellent general purpose bottom sampler.  In
 practice it operates better than either the Petersen
 or Franklin-Anderson over the full range of bottom
 types.  It can also obtain bottom samples with little
 or no disturbance and with the protecting screens
 removed or folded back, direct access can be had to
 the sediment surface of the sample.  Such access to
 an undisturbed sample makes it suitable for geochem-
 ical, sedimentological, biological, and structural
 studies.  Because of the large sample volume and
 its relatively undisturbed state, this sampler is
 very suitable for population studies of the bottom
 sediment fauna.
An excellent general purpose sampler, though perhaps
 a little heavy for small boat operation.  This
 sampler is capable of working with almost equal
 success on all types of bottom material.  It
 provides a sample even less disturbed than the
 Ponar, making it the most suitable sampler (under
 test) for detailed geological studies of the
 sediment surface.  The sampler volume is signif-
 icantly less than that of the Ponar, and the
 quantity of material sampled at maximum cutting
 depth is also less than the Ponar.  These two
 points may, therefore, favor the Ponar for certain
 biological (population) studies.  On the other
 hand, the rapid rotation of the Shipek bucket, as
 opposed to the much slower closure of the Ponar's
 jaws, may make it more suitable for sampling
 sediment containing a significant population of
 nonsessile forms.
                                  2-10

-------
'manager  is  responsible for making these decisions using the general
 guidance offered  in Section 1.
            The  responsibility of the field personnel is to implement
 a  collection program that minimizes sample contamination due to the
 process  of  sampling.  Sampling locations should be approached from
 the  downwind or downcurrent side to avoid effects due to the sampling
 vessel itself or  wastes from the vessel.   In addition, a sampling
 hierarchy should  be established to prevent undue sample contamination
 as a consequence  of the previous sampling effort.  Surface water
 samples  should  receive the highest priority, followed by water from
 increasing  depth.  The last samples to be obtained at each station are
 sediment samples  because of the potential for loss as they are
 retrieved through the water column.

                            Sample Handling

            The  method of sample handling after collection and prior to
 analysis is determined by the type of test to be run and the specific
 parameter being quantified.  There are three types of chemical tests
 being considered  in this manual:  standard elutriate test, bulk
 analysis, and elemental partitioning.  Each test has certain sample
 handling requirements that are summarized in schematic form in
 Figure 2-1.
            Each test provides different information on the sample and,
 therefore,  requires different handling.  The elutriate test indicates
 the  ability of  chemical constituents to migrate from the solid phase
 to the liquid phase.  '    Since chemical forms migrate differently,
 sample alterations due to drying, freezing, or air oxidation are to be
 avoided  and the test should be completed as soon as possible (preferably
 within 1 week of  collection).2  Elemental partitioning provides
 information on  the distribution of chemical constituents among
 several  defined sedimentary chemical phases.       Samples should be
 processed as quickly as possible and sample alterations due to drying,
 freezing, or air  oxidation are to be avoided.  The third test considered
 is bulk  analysis, which provides information on the total concentration

                                 2-11

-------
                                       2
of chemical constituents in the sample.    Since chemical speciation
is less important for this test, greater flexibility in sample storage
is acceptable with this test, provided the processes of freezing  and/
or drying do not cause the chemical degradation, or volatilization of
chemical contaminants of interest.
           An examination of Figure 2-1  reveals that all three tests
can be run on dredge, core, or sectioned core samples.   The difference
is that more stringent sample storage requirements must be met for the
elutriate test and elemental partitioning.  While there is greater
flexibility in storing bulk analysis samples, there is  less flexibility
in sample utility after the choice  is made.  That is, a sample stored
in a dried or frozen state can be analyzed for total content but  cannot
be used in the elutriate test or elemental partitioning, vhile a  sample
properly preserved for either of the latter tests, can also be used in
bulk analysis testing.  In addition, samples stored for the elutriate
test or elemental partitioning also can  be used for toxicity or bioassay
testing.  While guidance for toxicity testing is beyond the scope of
this manual, it is important to realize  that toxicity is influenced by
chemical form, in addition to other factors, and that certain storage
practices, such as sample drying and freezing, can alter the toxicity
of a sample.1**
           A sample to be used in an elutriate test or  a sediment
fractionation analysis should be stored  under conditions that provide
for original moisture, refrigeration, and minimum atmospheric contact.
However, a sample for bulk analysis may  be stored in a  wet, dried, or
frozen state.  The selection among  these options should be based
primarily on the specific chemical  analysis to be completed.  Guidance
for selection between the three methods  of bulk analysis sample
storage is summarized in Table 2-3.
           The longest list of parameters is associated with sediment
samples stored in a wet condition.   These conditions are most similar
to in situ conditions and, therefore, subject to a smaller force  to
produce a change.  Drying or freezing of a sample can alter the total
concentration of some constituents  as well as the chemical speciation
of others.  Parameters in this category  are biological  oxygen demand

                                2-12

-------
        I Dredge Sample I
        I Core Sample |
ELUTRIATE TEST
Use wet sediment, do not dry
or freeze. Process In 1 week.
^
r
       Prepare standard elutriate
                Filer
        I Process as water sample |

               TY_OF_COIiSJJJ_UJNj|
                                          I Section Core |
                                                        BULK ANALYSIS
                                                     Sediment treatment
                                                    and storage depends
                                                       on parameter.
                                                    I
                           Wet
                                        Freeze!
                                               \
                           SEDIMENT FRACTIONATION
        Use wet sediment, do not dry or freeze sample.   Begin as soon
        as possible.  Preserve anaerobic integrity of sample.	
          Res idue
           NH^OAc*
         Exchangeable

        | Exchangeable
Residue
Process must be conducted
under N2 atmosphere.
                                  Residue
                                                       Residue
                                        Moderately
                                        Reducible
                                                   HF + HN03 +
                                                   Fuming HNOs
                                                  [Residual    |

Figure 2-1.    Comparative methods of sediment sample handling
                             2-13

-------
                               Table 2-3
                Recommended Method of Sample Storage as
a Function of Bulk
Sediment Analyses to be Performed

Wet
CEC
Clz Demand
BOD
COD
SOD
Carbamates
PH
SRP
Redox
Total solids
Volatile solids
Sulfides
Phenoxy acids
Particle size
TOC.
TIG
Pesticides
Phenolics
Specific Gravity







NHa
N02~
N03~
Org-N
TKN
0 & G
PCB's
Org-P
Total-P
PAH
Hg
Al
As
Cd
Ca
Cr
Cu
Fe
Pb
Mg
Mn
Mo
Ni
Se
Zn
Dry
Particle size* Al
TOC As
TIC Cd
PCB's Ca
Pesticides Cr
Org-P Cu
Total-P Fe
PAH** Pb
Mercuryt Mg
Mn
Mo
Ni
Se
Zn











Freeze
Particle size* Al
TOC As
TIC Cd
0 & G Ca
PCB ' s Cr
Pesticides Cu
Phenolics Fe
Org-P Pb
Total-P Mg
PAH Mn
Mercury Mo
Ni
Se
Zn











*  Dispersed particle size probably not affected by drying or freezing.
  Apparent particle.size may be affected.
** PAH = polycyclic aromatic hydrocarbons.
t  Mercury may be lost if sample is dried at too high a temperature.

-------
(BOD), chemical oxygen demand (COD), sediment oxygen demand (SOD),  and
chlorine demand.  Since drying or freezing can result in sample oxi-
dation and, hence, a reduction of these parameters, these tests should
be run on wet samples.  Other variables that should "be quantified on
wet samples are cation exchange capacity (CEC), carbamates, pH, Eh
(redox potential), soluble reactive phosphate (SRP), total solids,
volatile solids, sulfides, and phenoxy acids.  The fact that analytical
results can be altered due to sample oxidation (affects sulfide, SOD,
COD, and Eh results), sample drying (CEC, pH), sample volatility
(phenols, volatile solids), and constituent instability (carbamates)
also would indicate that these constituents should be analyzed as soon
as possible after sample collection.
           Parameters that can be run on dried or frozen samples are
metals, stable organics, total nutrients, and minerals.  However, there
are precautions that should be considered.  For example, volatile
substances (Hg, Se, some phenolics and organics, and possibly ammonia)
may be lost if the sample is dried at too high a temperature.  Another
borderline parameter is particle size.  If a dispersed particle size is
to be determined, then drying or freezing should not affect the sample.
However, apparent particle size may be altered by these storage
procedures.

                          Sample Preservation

           It would be ideal if samples could be analyzed immediately.
However, this is seldom possible because of the number of samples to
be collected, the fact that all analytical equipment cannot be
satisfactorily operated in the field, and manpower limitations.
Therefore, samples should be treated when necessary with appropriate
preservatives to minimize chemical changes between the time of collec-
tion and the time of analysis.
           It cannot be stressed too strongly that a sample will be
subject to chemical, biological, and physical changes after collec-
tion.15  The use of sample preservatives does not halt these changes
but serves to slow or minimize them and, thus, provides more time to

                                2-15

-------
complete the required analyses.  Approaches that have been used to
preserve samples are refrigeration, pH adjustment, addition of
chemicals, and sample extraction.  The selection of appropriate
preservation techniques also depends on the specific analysis to be
conducted.  Recommended techniques for water sample preservation are
summarized in Table 2-h.
           It should be apparent that there is no universal preservative
and that a technique used to minimize the changes in one parameter may
alter the concentration or interfere in the analysis of another sub-
stance.16  This problem can be overcome by splitting the sample at the
time of collection.  Individual aliquots of the sample can then be
preserved as required without interfering with other analyses.  In
general, the number of subsamples that will be required will be equal
to the number of different preservatives that must be used.
           A similar listing of preservation techniques for specific
chemical constituents in sediment samples has not been prepared.  This
results from the fact that sediment preservatives have not been as
thoroughly evaluated as water sample preservation techniques.  Also,
the preservative would have to be thoroughly mixed with the sample,
which would result in the destruction of sample integrity.  However,
as a general rule, sample containers should be selected based on the
guidance provided for water samples (Table 2-U).  That is, aliquots
for metal or nutrient analysis may be stored in plastic or glass
containers; but aliquots for organic analysis should be stored in
glass containers with teflon-lined caps.  In addition, the following
approach(es) is (are) recommended, depending on the test to be
performed:
           a_.  Elutriate test.  Sediment samples should be stored wet,
               at UC, and in an airtight container.2  The elutriate
               test procedure should be initiated within 1 week of
               sample collection.  The standard elutriate that results
               from this procedure can then be analyzed immediately or
               treated as a water sample and split and preserved, as
               discussed earlier.
           b_.  Sediment fractionation.  Sediment samples scheduled for
               fractionation analysis should also be stored wet, at
               ^C, and in an airtight container.  The last requirement
                                 2-16

-------
                               Table 2-k
           Recommended Water Sample Preservation Techniques
15
Parameter
Sample
Volume
Container* ml
Total organic carbon P
Total inorganic carbon P
Chlorine demand
Aluminum
Arsenic
Cadmium
Calcium
Chromium
Copper
Iron
Lead
Magnesium
Manganese
Mercury
Molybdenum
Nickel
Selenium
Zinc
Ammonia-nitrogen
Nitrate-nitrogen

P
P
P
P
P
P
P
P
P
P
P

P
P
P
P
P
P

, G
, G
, G
, G
, G
, G
, G
, G
, G
, G
, G
, G
G
, G
, G
, G
, G
, G
, G

100
100
Preservative
H2S04 to pH <
UC
Air Seal
2
Storage
Time
2U -
None
100-200t

100-200+
100-200t
100-200t
100-200t
100-200t
100-200t
loot
100-200+
500
100-200t
100-200+

100-200+


(Continued)
HN03
HNOs
HN03
HNOs
HNOs
HNOs
HNOs
HN03
HN03
HN03
HNOs
HNOs
HNOs
HNOs
HNOs
H2SO,
H2SO>*

to
to
to
to
to
to
to
to
to
to
to
to
to
to
to
to
to

pH <
pH <
pH <
pH <
pH <
pH <
pH <
pH <
pH <
pH <
pH <
pH <
pH <
pH <
pH <
pH <
C
pH <
C

2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2
2

6
6
6
6
6
6
6
6
6
6
2
6
6
90
6
2l
48 hr
0**
0
mo
mo
mo
mo
mo
mo
mo
mo
mo
mo
wk
mo
mo
days
mo
4 hr
2U hr


*  P = plastic;  G = glass.
** One reference indicates  TIC may be preserved  for  3 mo  in a  sealed
    bottle with  HgCl2  (W. S.  Wong.   Deep Sea Research 17:9-17  (.1970).
t  Sample can be used  for other metal analyses.
                                                        (Sheet 1 of 3)
                                 2-17

-------
Table 2-h (Continued)
Sample
Volume
Parameter Container ml-8,
Nitrite-Nitrogen P, G
Organic -Nitrogen P, G
Total Kjeldahl
Nitrogen P, G

Oil and grease G

Biochemical dxygen
demand P, G 300 ml-2 %
Chemical oxygen demand P, G 200 ml

PCB's G 2.1
Organochlorine pesti-
cides G 1 H
Chlorinated phenoxy
acid herbicides G 1 &

Organophosphates and
carbamates G 1 H

Phenolic s G 500 ml-1 *


Soluble reactive
phosphates P, G

Organic phosphate
Total phosphorus P, G
Redox potential P, G 100 ml
pH P, G 100 ml

Preservative
H2S04 to pH < 2
H2 S04 to pH < 2

H2 SQ, to pH < 2
hoc
H2S(\ or HC1
to pH < 2

^C
HzSOi to pH < 2
hC
hC

hC

HaSOif to pH < 2
hC

H2SOii to pH < 3
10 g Na2SOi4
!, 0.1-1.0 g CuSOit
H3POit to pH < h
hc

Filter
hc

hC
None
None
hC
Storage
Time
2h hr
2h hr

2h hr

2h hr


6 hr
7 days










2h hr




2h hr

7 days
None
6 hr

      (Continued)





        2-18
(Sheet  2 of 3)

-------
                           Table 2-k (Concluded)
Parameter
Total solids
Volatile solids
Sulfi.des
Polynucleated aromatic
hydrocarbons
Container
P, G
P, G
P, G
G
Sample
Volume
ml Preservative
200tt ^C
200tt kC
2 ml ZnOAc

Storage
Time
1 days
1 days
2k hr

tt Sample can be used for total solids  and volatile  solids.
                                                          (Sheet  3 of 3)
                                   2-19

-------
               is especially important since the first  two steps  of
               the fractionation procedure must also be carried out
               under a nitrogen atmosphere. "*  Fractionation procedures
               should commence as soon as possible and, preferably, no
               later than 1 week after sample collection.   As the
               operationally defined fractions are prepared, they may
               be analyzed immediately or preserved for specific
               constituents and analyzed at a later date.

           c_.  Bulk analysis.  There- is more flexibility in selecting a
               storage technique for samples to be analyzed for total
               concentration.  The reason is that, unlike the previous
               two procedures which measure specific forms of chemicals
               in the sediments, bulk analysis measures the total content
               of the sample.  Therefore, the results are not affected
               by processes such as oxidation and air drying that alter
               the species distribution of chemicals in the samples.
               The choice between storing a sample in a wet condition,
               dried condition, or a frozen condition should be based
               largely on the analyses to be run as illustrated in
               Table 2-3 and discussed previously.

           d.  Tissue analysis.  There are two factors  that must  be
               considered when it is necessary to store biological
               tissue.  The first factor is the stability of the
               specific chemical entity.  The second factor is the
               stability of the tissue itself since analytical results
               are expressed on a wet weight and/or a dry weight  basis.
               Thus, concentrations can be altered if the tissue  is
               dehydrated during storage even though the chemical of
               interest may be stable.  A suggested approach would be
               to record the appropriate wet and/or dry weights of
               the tissue samples as soon as possible.   Tissue samples
               should be digested (for inorganic constituents) or
               extracted (for organic constituents) and the digest
               or extract can then be stored according  to the guidance
               presented in Table 2-k.
                            Quality Control


Objectives
           An integral part of any sampling program must be a quality

assurance or control program.  The objectives of this program should
be to determine the quality of the data (accuracy and precision) and
to control the quality of the data' (variability).  The responsibility

for this program must be shared between the field and laboratory
personnel since quality control begins with sample collection and not
                               2-20

-------
sample analysis.    It is important that field personnel be made aware
of the fact that the greatest potential for sample contamination occurs
during the preanalysis steps of sample collection, handling, preserva-
tion, and storage.
           There are several functions that can be performed by field
personnel to assist in an overall quality control program.  These
include:
           EU  Providing a representative sample for analysis.
           b_.  Providing replicate samples to define variation at a
               single point.
           c_.  Providing a sufficient amount of sample to allow
               detection.
           d_.  Spiking occasional samples to correct for sample decay
               between collection and analysis.
           _e.  Initiating analysis or appropriate storage procedures
               immediately after collection.
           _f_.  Properly labeling and recording the dates and location
               of sample collection.
           The need to include all of these functions will depend on
the specific purpose of the project.  As discussed in Section 1,
representativeness is a difficult property to assess.  Also, replicate
sampling at each location may be desirable but not essential for all
projects.  However, collection of sufficient sample, utilization of
appropriate storage procedures, and proper sample identification
should be an integral part of every sample collection effort.
           Laboratory personnel also have certain functions that must
be satisfied in order to complete the quality control program.17  Some
of the more obvious duties are:
           ii.  Using acceptable techniques for analysis.
           b_.  Completing the analysis immediately (ideally) or
               within prescribed storage limits that are parameter
               specific.
           c_.  Performing replicate analysis on approximately 5 to 10
               percent of samples processed.
           d_.  Adding standard solution spikes to approximately 5 to 10
               percent of samples processed and determining recovery..
           e_.  Using an internal laboratory standard to check perfor-
               mance of analytical method.

                                 2-21

-------
           f_.  Analyzing externally prepared reference and performance
               (unknown) samples on a routine "basis.
A more detailed list of ideal quality control activities that was
prepared by Delfino18 is presented in Table 2-5.
Work load
           It is suggested that a quality control program should consist
of approximately 15 to 20 percent of the total analytical work load.
This should consist of the following:
           a_.  Five percent duplicate sample collection.
           b_.  Five percent replicate determinations.
           c_.  Five percent spike recovery.
           (1.  Five percent external reference, field blanks, or
               unknown samples and sample splitting with other
               laboratories.
These are discussed in more detail in the following paragraphs.
           Duplicate samples.  At stations selected at random, duplicate
samples are collected from two sets of field equipment installed at the
site, or duplicate grab samples are collected.  This provides a check
of sampling equipment and technique for precision.
           Split samples.  A collected sample is split and each
aliquot is analyzed as an independent sample.  The samples may be
reanalyzed by the same laboratory or analyzed by two different
laboratories as a check of the analytical procedures.
           Spiked samples.  Known amounts of a particular constituent
are added to an actual sample or to blanks of deionized water at
concentrations at which the accuracy of the test method is satisfactory.
The amount added should be coordinated with the laboratory.  This
method provides a proficiency check for accuracy of the analytical
procedures.
           Sample preservation blanks.  Acids and chemical preserva-
tives can become contaminated after a period of use in the field.
The sampler should add the same quantity of preservative to some
distilled water as normally would be added to a water or sediment
sample.  This preservative blank is sent to the laboratory for analysis
of the same parameters that are measured in the sample and values for
                                2-22

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                               Table 2-5
               Ideal Control Activities for Documenting
                   the Validity of Laboratory Data*
Verify calibration curves.
Confirm instrumental calibrations (wavelength, temperatures, etc.).

Monitor precision by performing replicate analyses on ca. 5 to 10
 percent of  samples processed.
Perform multiple replicates during the day and compute standard devia-
 tion; if possible, compute relative standard deviation when dealing
 with wide ranges of concentrations.
Document recovery by adding standard solution spikes to ca. 5 to 10
 percent of  samples processed; determine percent recovery of spikes.
Use an internal laboratory standard to trace performance on a given
 analytical method; match matrix as closely as possible to sample(s).

Split samples with other laboratories performing similar analyses, at
 least on a  quarterly basis.
Analyze an externally prepared performance sample [e.g. EPA, National
 Bureau of Standards (NBS), etc.] at least quarterly.
Analyze an externally prepared performance sample (unknown) from EPA
 or other source at least annually.
Develop quality control charts for precision and recovery performance;
 use charts  to monitor daily laboratory performance; generate with a
 computerized data handling system, if possible.
Develop correlation data between analyses of similar meaning and use
 as cross-checks on validity of results, e.g., conductivity and total
 dissolved solids, turbidity and total suspended solids, TOC** and
 ;BOD/COD, equivalent charge balance.
Calibrate analytical balances when irreproducibility is noted; service
 balances on an annual basis.
Rotate chemical inventory to eliminate older chemicals and reagents.
Develop replacement schedule for standard solutions; discard sooner if
 calibration curves change and this cannot be related to instrumental
 variation.
Develop instrument maintenance records; enter all service, adjustments,
 etc., including problem diagnosis and resolution; state if data were
 reported when instrument was out of calibration and, if so, explain
                              (Continued)

                                                                  I Q
' * Perform on a daily basis unless noted otherwise; after Delfino.
 ** TOC = total organic carbon.

                                 2-23

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                         Table 2-5 (Concluded)
 disposition of 
-------
the blank are then subtracted from the sample values.   Liquid chemical
preservatives should be changed every 2 weeks or sooner if contamination
increases above predetermined levels.
           It is also essential that reagent blanks or distilled water
and solvents used in the laboratory be routinely analyzed for contami-
nation.  These values are also subtracted from the determined sample
values.  Reagents, solvents, and distilled water should be purified
or replaced if contamination exceeds predetermined levels.
Present limitations
           The theory of quality control was developed for an industrial
application approximately 50 years ago to evaluate the quality of a
product.  Unfortunately, there are differences between environmental
samples and industrial produc-ts that make it difficult to satisfactorily
apply industrial quality control techniques.  Five major components
that contribute to total error in an environmental sample are:  site
selection, sampling, measurement method, reference sample, and data
handling error.19  The largest error, particularly for sediment
samples, is site selection, over which there is no control.  In
addition, quality control statistics are dependent upon the true
sample value being known, which is not generally the case with
environmental samples.  Since there is no expected value for a
randomly selected sample, much more reliance must be placed on
standard and spike recovery, and replicate determinations, to
indirectly evaluate the accuracy of testing methodology.17

                        Types of Chemical Tests

           Three types of chemical tests are considered for analysis
of dredged and/or fill material samples:  standard elutriate test,
sediment fractionation, and bulk or total analysis.  The selection of
any one of these tests or combination of these tests should be based
on the purpose of the study as discussed in Section 1.
           The elutriate test is a short-term, sediment-leaching
procedure.  It consists of agitating a known volume of sediments/fill
                               2-25

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material with a known volume of site water.   The suspension is then
filtered and the filtrate analyzed.   Thus, the test provides an indica-
tion of the chemical constituents likely to be released to the water
column during a disposal/filling operation.2'20'21   Since the sediment-
to-liquid ratio used in the test is  based on hydraulic dredging ratios,
results from the elutriate test will probably overestimate the release
from less dynamic dredging techniques such as hopper or clamshell
dredging.
           The purpose of the elutriate test is to  provide information
on the potential effects of a disposal operation on water quality.
Results can then either be used to estimate the extent of a resource
that will be influenced by the proposed discharge or used to compare
the results to appropriate water quality criteria.
           The first option would be preferred because it provides
information on the amount of the receiving water necessary to assimilate
the proposed discharge and whether other critical uses such as spawning
grounds or water intakes may be impacted.  This is  accomplished by
determining whether the required mixing zone overlaps with other areas
                      n
of specific water use.
           The second option is less desirable for  two reasons.  First,
a comparison of elutriate test concentrations with  criteria would be
                                                          2 2
overly conservative because site dilution is not included.    Second,
water quality criteria have an implied exposure time ranging from
96 hr to many months, while dredged material perturbations persist
for 30 min to 2 hr.  Since disposal plumes exist for shorter time
periods, a direct comparison to criteria would be even more conserva-
tive.  Because of the nature of the  comparisons, an elutriate test
result less than established criteria would indicate that adverse
water quality impacts would not be expected.  However, an elutriate
test result exceeding established criteria would not necessarily
imply that adverse water quality impacts would occur.
           Elemental partitioning or sedimentation  fractionation
studies are the most complex of the tests considered in this manual.
They have only been used in studies  involved with metals and nutrients
(carbon, nitrogen, phosphorus) to date.10 12'23  The procedure
                                2-26

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consists of exposing the sample to a series of leaching agents of
increasing strength.  The first step consists of centrifugation/
filtration of the sample to isolate interstitial water.  The solid
residue is then sequentially leached with ammonium acetate,  hydroxy-
lamine, hydrogen peroxide, dithionate, and a hydrofluoric acid-nitric
acid mixture.10 12  Results provide an indication of the distribution
of chemicals in sediments and fill material and the harshness required
to mobilize that constituent.11 13
      93.  At this time, there is no simple or universal method of
evaluating fractionation studies.  When fractionation results were
correlated with elutriate test results,11 the highest degree of corre-
lation occurred between elutriate concentrations and the interstitial
and exchangeable phases.  This suggests that the elutriate test is a
measure of the most mobile sediment constituents.  In another study,
no relationships were observed between chemical fractionation results
                                9 o
and biological uptake of metals.    A third study correlating long-term
release with fractionation results demonstrated the highest  degree of
correlation with elutriate and interstitial water concentrations.l  The
number of correlations decreased markedly as the strength of the extract
increased.  Results demonstrate the complexity of evaluating the poten-
tial effects of chemicals in sediments and suggest that the  more tightly
bound substances are less likely to create environmental problems.
      9^.  Bulk analysis results provide information on the  total
concentration of chemical constituents in the samples being  analyzed.
The procedure consists of a strong acid digest or an organic solvent
extraction of the sample.  Because specific chemical forms or chemical
distribution are not of importance with this test, bulk analysis
generally allows more flexibility in sample handling.
      95.  Total analysis results can be used to calculate the mass
(concentration x volume) of a specific constituent involved in a
dredging/filling operation.  Results also can be used for a crude
comparison of sediments and/or fill material with the proposed disposal
site.  However, it is recommended that bulk analysis results not be used
to evaluate potential environmental impacts of a proposed disposal
operation.  References cited earlier demonstrated little, if any,

                                 2-27

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relationships between total sediment concentration and biological
uptake or changes in water quality.   Also, a review of the technical
literature indicated no correlations between total composition and
                                     2 4
sedimentary effects on water quality.
Elutriate test
           The elutriate test is a simplified simulation of the
dredging and disposal process wherein predetermined amounts of
dredging site water and sediment are mixed together to approximate
a dredged material slurry.2'20  The elutriate in the supernatant
resulting from the vigorous 30-min shaking of one part sediment from
the dredging site with four parts water (vol/vol) collected from
the dredging site followed by a 1-hr settling time and appropriate
centrifugation and 0.1*5 y filtration.  Thus, it will be necessary to
collect both water and sediment samples to perform the elutriate test.
When evaluating a dredging operation, the sediment should be collected
at the dredging site and the water should be collected at the dredging
and the disposal site.  To evaluate a fill material activity, samples
should be collected from the source of the fill material and the water
should be collected from the disposal site.
           Water sample collection.   Collection should be made with an
appropriate noncontaminating water sampling device.  Either discrete
samplers such as Kemmerer or Van Dorn samplers or continuous collectors
such as submersible pumps may be used.  The volume of water required
will depend on the number of analyses to be performed.  For each sample
to be subjected to elutriate testing, it is suggested that a minimum of
U  be collected at the disposal site and 8  be collected at the
dredging site to evaluate, a dredging operation and/or 12 H be collected
at the disposal site to evaluate a fill material disposal operation.
This will provide k  of water for analyses and sufficient water to
prepare triplicate 3-& elutriates.  (Each elutriate should yield
2.0 to 2.2 H of standard elutriate for analysis.)  If the samples are
to be analyzed for trace organics or a large number of constituents,
a proportionately larger initial sample should be collected.
           Samples must be stored in glass containers if trace
organic analyses are to be performed.  Generally, either plastic or
                                2-28

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glass containers may be used for other parameters.   The samples should
be maintained at kC until analyzed but never frozen.  The storage
period should be as short as possible to minimize changes in the
characteristics of the water.  Disposal site water should be analyzed
or split and preserved immediately.  The remainder of the water should
be used in the elutriate test, which should be processed within 1 week
of collection.
           Sediment sample collection.  Samples should be taken from
the fill or the dredging site with a grab or a corer.  Approximately 3 &
of sediment or fill material would provide sufficient sample to prepare
triplicate 3-& elutriates.  Again, if the resultant standard elutriates
are to be analyzed for trace organics or a large number of constituents,
a proportionately larger initial sample should be collected.
           Samples may be stored in plastic bags, jars, or glass
containers.  However, if trace organic analyses are to be performed,
glass containers with teflon-lined lids are required.  A special
precaution that must be taken with sediment samples is to ensure
that the containers are completely filled with sample and that air
bubbles are not trapped in the container.  This step is necessary to
minimize sample oxidation that could influence elutriate test
results.2'22
           The samples should be stored immediately at UC.  They must
not be frozen or dried prior to use.  The storage period should be as
short as possible to minimize changes in the characteristics of the
sediment.  It is recommended that samples be processed within 1 week
of collection.
           Apparatus.  The following apparatus are required to perform
the elutriate test.  Prior to use, all glassware, filtration equipment,
and filters should be washed with 5 to 10 percent (or stronger)
hydrochloric acid (HCl) and then rinsed thoroughly with deionized
water.  The necessary apparatus include:
           i.  Acid-rinsed plastic bottles for collection of water
               samples.
           b_.  Plastic qars pr bags ("Whirl-Pak," plastic freezer
               containers, etc.) for collecting dredged or fill
               material samples.
                                2-29

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c_.  Laboratory shaker capable of shaking 2-& flasks at
    approximately 100 excursions/minute.  Box type or
    wrist-action shakers are acceptable.

d:.  Several 1-& graduated cylinders.

e_.  Large (15 cm) powder funnels.

f_.  Several 2-., large-mouth graduated Erlenmeyer flasks.

g__.  Vacuum or pressure filtration equipment, including
    vacuum pump or compressed air source, and an appropriate
    filter holder capable of accomodating UT-, 105-, or
    155-m-diameter filters.

h_.  Membrane filters with a 0.^5-V pore-size diameter.
    The filters should be soaked in 5 M HC1 for at least
    2 hr prior to use.

i_.  Centrifuge capable- of handling six 1- or 0.5-& centri-
    fuge bottles at 3000 to 5000 rpm.  International Model
    K or Sorval Super Speed are acceptable models.

j_.  Wide-mouth, 1-gal capacity glass jars with teflon-
    lined screw-top lids for use as sample containers
    when samples are to be analyzed for trace organics.
    (It may be necessary to purchase jars and teflon
    sheets separately; in this case, the teflon lid
    liners may be prepared by the laboratory personnel.)

Test procedure.  The stepwise test procedure is given below:

&_.  Subsample a minimum volume of 1 H each of dredging site
    and disposal site water.  If it is known in advance
    that a large number of measurements are to be performed,
    the size of each subsample should be increased to meet
    the anticipated needs.

b_.  Filter an appropriate portion of the disposal site
    water through an acid-soaked O.H5-U pore-size membrane
    filter that has been prerinsed with approximately
    100 ml of disposal site water.  The filtrate from the
    rinsing procedure should be discarded.
c_.  Analyze the filtered disposal site sample as soon as
    possible.  If necessary, the samples may be stored at
    kC after splitting and the appropriate preservatives
    have been added (Table 2-U).  Filtered water samples
    may also be frozen with no apparent destruction of
    sample integrity.

d_.  Repeat steps a^, b_, and c_ with dredging site water.
    This step is omitted with a fill material sample.
e_.  Subsample approximately 1  of sediment from the well-
    mixed original sample.  Mix the sediments and
    unfiltered dredging site water in a volumetric
    sediment-to-water ratio of 1:U at room temperature
    (22 +_ 2C).  This is best done by the method of

                   2-30

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                volumetric displacement.23  One hundred mililiters of
                unfiltered dredging site water is placed into a
                graduated Erlenmeyer flask.   The sediment subsample is
                then carefully added via a powder funnel to obtain a
                total volume of 300 ml.   (A 200-ml volume of sediment
                will now be in the flask.)  The flask is then filled
                to the 1000-ml mark with unfiltered dredging site
                water, which produces a slurry with a final ratio of
                one volume sediment to four volumes water.

            This method should provide TOO to 800 ml of water for

analysis.  If the analyses to be run require a larger volume of water,

the initial volumes used to prepare the elutriate slurry may be
proportionately increased as long as the solid-to-liquid ratio remains

constant (e.g. mix hOO ml sediment and 1600 ml unfiltered dredging

site water).  Alternately, several 1-& sediment/dredging site water

slurries may be prepared as outlined above and the filtrates combined

to provide sufficient water for analysis.  The procedure continues as

follows:
            f_.  (l)  Cap the flask tightly with a noncontaminating
                     stopper and shake vigorously on an automatic
                     shaker at about 100 excursions per minute for
                     30 min.  A polyfilm-covered rubber stopper is
                     acceptable for minimum contamination.

                (2)  During the mixing step given above, the
                     oxygen demand of the dredged material may cause
                     the dissolved oxygen concentration in the elutriate
                     to be reduced to zero.  This change can alter
                     the release of chemical contaminants from dredged
                     material to the disposal site water and reduce
                                                               21
                     the reproducibility of the elutriate test.
                     If it is known that anoxic conditions (zero
                     dissolved oxygen) will not occur at the disposal
                     site or if reproducibility of the elutriate test
                     is a potential problem, the mixing may be
                     accomplished by using a compressed air-mixing*
                     procedure instead of the mechanical mixing
                     described in Step f_  (l).  After preparation of
                     the elutriate slurry, an air-diffuser tube is
                     inserted almost to the bottom of the flask.
                     Compressed air should be passed through a
                     deionized water trap and then through the
                     diffuser tube and the slurry.  The flow rate
                     should be adjusted to agitate the mixture

* This procedure can cause the loss of highly volatile chemical con-
  stituents.  If volatile Materials are of concern, compressed air
  mixing should not be used.

                                2-31

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                     vigorously for 30 min.   In addition,  the flasks
                     should be stirred manually at 10-min  intervals
                     to ensure complete mixing.

            g_.        After 30 min of shaking or mixing with air,
                     allow the suspension to settle for 1  hr.

            h_.        After settling, carefully decant the  supernatant
                     into appropriate centrifuge bottles and then
                     centrifuge.  The time and revolutions per minute
                     during centrifugation should be selected to  reduce
                     the suspended solids concentration substantially
                     and, therefore, shorten the final filtration
                     process.  After centrifugation, vacuum or pressure
                     filter approximately 100 ml of sample through a
                     O.U5-V membrane filter and discard the filtrate.
                     Filter the remainder of the sample to give a
                     clear final solution (the standard elutriate) and
                     store at ^C in a clean, noncontaminating container
                     in the dark.  The filtration process  is intended
                     for use when the standard elutriate is to be analyzed
                     for conventional chemical contaminants.  When the
                     elutriate is to be analyzed for organic contaminants
                     and PCB's, filtration should not be used since
                     organic concentrations can be reduced by sorption.
                     Centrifugation should be used to remove particulate
                     matter when the standard elutriate is to be
                     analyzed for specific organics.
            i_.        Analyze the standard elutriate as soon as possible.
                     If necessary, the samples may be stored at UC
                     after splitting and the appropriate preservatives
                     have been added.
            j_.        Prepare and analyze the elutriate in  triplicate.
                     The average of the three replicates should be
                     reported as the concentration of the  standard
                     elutriate.
Sediment fractionation
            Chemical constituents associated with sediments may be

distributed in many chemical forms.  The purpose of a fractionation
procedure is to better define this distribution.  This objective  is
achieved by leaching a sample with a series of successively harsher
extraction agents.  Reagents used in the procedure to be described
below consist of interstitial water, ammonium acetate, hydroxylamine,
hydrogen peroxide, citrate-dithionate, and hydrofluoric acid-nitric

acid.
            The premise of the fractionation procedure is  that a

specific geochemical phase is defined by a specific chemical

                                2-32

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extraction agent.  Thus, the ammonium acetate extract is referred to as
the exchangeable phase, and the citrate-dithionate extract is referred
to as the moderately reducible phase.  These relationships have not been
rigorously demonstrated and, therefore, the fractions are only opera-
tionally defined.
            The use of fractionation results at this time appears to be
limited to supporting other studies.  That is, results have greater
value in sediment research studies than in the regulatory decision-
making process.  Limited results vith sediment fractionation data
suggest a higher correlation with the more labile sediment phases
(interstitial water, exchangeable phase) and elutriate test results11
and long-term water quality changes.10
            A major limitation of the fractionation procedure is that
previous experience is limited to heavy metals and nutrients.  A broad
spectrum analysis of the individual fractions has been limited by small
sample size, particularly the interstitial water fraction.
            Sample collection.  Samples should be collected with a grab
or a corer.  Because the distribution of sediment-associated chemicals
can be altered by processes such as drying and oxidation, samples should
be kept wet and exposure to the atmosphere should be minimized.  Samples
collected with a grab or dredge must be quickly transferred to a con-
tainer and air bubbles must be excluded from the container.  Corer
samples should be sealed in the core liner and returned to the labora-
tory in an upright position.  Previous studies have shown that a 15-cm
section from a 7-5-cm-diameter core can provide sufficient material to
perform fractionation studies for eight metals and four nutrients.11
            The samples should be stored immediately at hC.  They
must not be frozen or dried prior to use.  The storage period should
be as short as possible to minimize changes in the distribution of
chemical constituents in the sediments.  It is recommended that samples
be processed within 1 week of collection.
            Apparatus.  The following apparatus is required to perform
the elemental partitioning procedure.  Prior to use, all glassware,
filtration equipment, and filters should be washed with 5 to 10 percent
HC1 and then rinsed thoroughly with deionized water.  This list of

                               2-33

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equipment includes:
            au  Plastic jars or bags ("Whirl-Pak," plastic freezer
                containers, etc.) for collecting grab or dredge
                samples or polyethylene liners for the collection of
                core samples.
            b_.  Glove box or disposable glove bag.
            c_.  Polarographic oxygen analyzer or alternate method to
                confirm the absence of oxygen in glove bag.
            cl.  Utensils for splitting cores and handling samples in the
                glove bag.
            e_.  250-ml and 500-ml polycarbonate centrifuge bottles.
            f_.  Refrigerated centrifuge.
            g_.  Vacuum filtration apparatus.
            hu  150-ml and 120-ml polyethylene storage bottles.
            i_.  Blender or porcelain mortar and pestle.
            j_.  Top-loading balance.
            k_.  Weighing dishes.
            !_.  Digestion block or hot plate.
            m.  Teflon beakers.
            ri.  50-ml volumetric flasks.
            Test procedure.  A step-wise sediment fractionation procedure
is given in the following paragraphs.
            To begin the procedure, prepare a glove box or disposable
glove bag.  Flush the system with nitrogen gas and maintain a positive
pressure nitrogen atmosphere.  Oxygen-free conditions in the glove bag
or box should be verified with a polarographic oxygen analyzer prior
to sample processing.  Initial sample handling and all steps in the
interstitial water and ammonium acetate extractions should be conducted
under a nitrogen atmosphere.
            Acid wash all hardware to be used in the extractions in
6 N_ HCL and thoroughly rinse with distilled water to minimize sample
contamination during processing.
            The initial separation removes the interstitial water phase.
This is accomplished by first placing the sealed sediment sample in the
glove bag.  After reestablishing the nitrogen atmosphere, extrude the
sediment core from its liner into a flat plastic container.  If the
core is to be sectioned vertically, 15-cm sections of a 7.5-cm-diameter
                                2-3**

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core have been shown to provide sufficient material for the sequential
fractionation procedure.  Each core section should "be split into halves
with one half (approximately 300 cc) being used for the interstitial
water testing and the remaining half used for all other analyses.  Place
the half section for the interstitial water analysis in an oxygen-free,
polycarbonate 500-ml centrifuge bottle in the glove bag and seal.
Centrifuge the sample in a refrigerated centrifuge (UC) at 900
revolutions per minute (^13,000 x g) for 5 min.  This should be
sufficient to recover Uo percent of the total sediment water.  After
centrifugation, return the sample to the glove bag and vacuum filter
the interstitial water through a oA5-y pore-size membrane filter.
Transfer the filtered sample to an acid-washed polyethylene bottle and
acidify to pH 1 with HC1 for preservation.
            If the sediment sample is not to be sectioned vertically,
decant excess water and blend the core or dredge sample.  Place
approximately 300 cc of the blended sample in an oxygen-free, poly-
carbonate 500-ml centrifuge bottle and seal.  Centrifuge this sample
for 5 min at 900 revolutions per minute (13,000 x g) in a refrigerated
centrifuge (kC) and then filter through a 0.^5-M pore-size membrane
filter under a nitrogen atmosphere.  The filtered sample may be
analyzed immediately or split for preservation and storage.
            The exchangeable phase is determined on the unused half of
the wet sediment sample that was blended for interstitial water
analysis.  Blend the wet sediment with an electrically driven poly-
ethylene stirrer contained in the glove bag.  Remove a subsample of
the homogenized sediment sample (blended core section, core, or grab
sample) for percent solids determination.
            Weigh a second subsample (approximately 20 g dry weight
of each homogenized sediment section into an oxygen-free, tarred,
250-ml centrifuge tube containing 100 ml deoxygenated 1 IJ ammonium
acetate, producing a suspension with an approximate solid-to-liquid
ratio of 1:5-  Adjust the pH of the acetate solution to the pH of the
surface sediments.  Seal the samples and then place on a wrist-action
shaker for 1 hr.  Centrifuge the samples at 6000 revolutions per minute
for 5 min and return them to the glove bag for further processing under

                                 2-35

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the nitrogen atmosphere.  Filter the sample through O.U5-y pore-size
membrane filters, retaining both the filtrate and the solid residue.
            The filtrate may be analyzed immediately or split and
preserved for specific constituents, as discussed for water samples.
This extract will also include the interstitial water components
since a fresh blended sediment sample was used.  Therefore, measured
concentrations should be reduced to compensate for the interstitial
water.  This can be accomplished as:
                                                                     (3)
            Wt exchangeable material = (Vol ext) (Cone ext) -
                    (Wt Sample) (l-g Solids) ,    *
                        (density water)
where
(Vol ext)       = volume of ammonium acetate.extract
(Cone ext)      = analytical concentration in ammonium acetate extract
(Wt Sample)     = wet weight of sample for ammonium acetate extraction
% Solids        = percent solids in sample
(density water) = density of water at temperature of sample
(IWC)           = interstitial water concentration of sample
            The easily^ reducible phase is performed with the solid
residue from the exchangeable phase determination.   This step and all
subsequent steps in the fractionation procedure can be conducted outside
the glove bag.  Add 50 ml Na sparged distilled- deionized water to the
centrifuge tube containing the solid residue from the 1 N_ ammonium
acetate extraction.  Agitate the sample with a stainless steel spatula
or a glass-stirring rod to ensure good washing efficiency.  Centrifuge
the suspension of 6000 revolutions per minute and discard the liquid
phase.  A portion of the solid residue will have to be set aside at
this point for a redetermination of percent solids.
            Blend the remaining sediment residue and transfer a 2-g
(dry weight equivalent) subsample to a 250-ml Erlenmeyer flask.  Add
100 ml of 0.1 M hydroxylamine hydrochloride-0.01 M nitric acid solution.
The resultant suspension will have a solid-to-extractant ratio of
approximately 1:50.  Seal the sample and place the suspension on a
wrist-action shaker (or equivalent) for 30 min.  Centrifuge the sample

                                 2-36

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at 6000 revolutions per minute for 5 min.   Decant and filter the liquid
phase through 0.1+5-y pore-size membrane filters.  The filtrate may be
treated as a water sample and analyzed immediately or split and preserved
for specific constituents.
            The solid residue is used in the organic and sulfide phase
extraction.  Wash the residue from the easily reducible'phase with 50 ml
distilled water,,  After agitating the suspension, centrifuge the sample
at 6000 revolutions per minute for 5 min and discard the supernate.
Subsample the residue for a percent solids determination so the organic
and sulfide results can be expressed on a dry weight basis.  Add 50 ml
of 30 percent hydrogen peroxide to the washed residue and adjust the pH
to 2.5 with HC1.  (The purpose of the pH adjustment is to prevent any
released metals from precipitating.)  Digest the sample at 95C for
6 to 8 hr.  Add 100 ml of 1 N. ammonium acetate buffered at pH 2.5 to
the digestate and shake for 1 hr.  Centrifuge the sample at 6000 revo-
lutions per minute for 5 min and filter the sample through 0.^5-y
pore-size membrane filters.  Treat the filtrate as a water sample
and analyze immediately or split and preserve as required.  Retain
the solid residue.
            The next fraction in the sequence is the moderately
reducible phase.  Wash the organic and sulfide phase solid residue
with 50 ml of distilled water; centrifuge as described earlier; and
discard the supernate.  Redetermine percent solids on a subsample
of the residue.  Add 100 ml of a citrate-dithionate solution
(l6 g sodium citrate + 1.67 g sodium dithionate/100 ml distilled
water) and mechanically shake the suspension for 17 hr.  Centrifuge
the sample at 6000 revolutions per minute for 5 min and filter the
supernate through a O.U5-V pore-size membrane filter.  Analyze the
filtered sample for moderately reducible constituents and retain
the solid residue for further treatment.
            The sample for residual phase digestion is obtained by
washing the moderately reducible phase residue with 50 ml of distilled
water; centrifuging at 6000 revolutions per minute for 5 min; and
discarding the supernate.  Dry the residue at 105C and transfer
                               2-37

-------
a 0.5-g dry weight subsample to a teflon beaker.  Add  15 ml hydro-
fluoric acid and 10 ml concentrated nitric acid;  cover the beaker;
and digest at 1T5C.  After evaporation to near dryness,  add  8 ml
fuming nitric acid stepwise in 2-ml increments.  Continue  evaporation
to near dryness.  Add 6 N_ HC1 to dissolve the residue, heating if
necessary.  Quantitatively transfer the solution to a  50-ml volumetric
flask and dilute to volume.  Analyze the sample immediately or preserve
subsamples for specific constituents.
            A schematic flow diagram for the fractionation procedure
is presented in Figure 2-2.  When this procedure is used,  a built-in
quality control check is to total each of the operationally defined
phases and compare to a total digest of the sample. The  data should
be considered suspect if they differ by more than 5 to 10  percent.
Bulk analysis
            A bulk analysis provides a measure of the  total concen-
tration of a specific constituent in the sample being  analyzed.  This
is accomplished by subjecting a sample to strong oxidation, acid
digestion, or organic solvent extraction.  The procedure  is similar
to that used for the residual phase digestion in the elemental
partitioning procedure discussed earlier.  Total sediment  concentrations
can be used to compare different sites and to identify major  point
sources.  However, because of the harshness of the extraction pro-
cedure, information on chemical distribution and/or potential
environmental impact is lost.
            Sample collection.  Samples for total analyses may be
collected with a dredge or grab sampler or a core sampler. Approxi-
mately 1 to 2 H of sediment or fill material should be taken  from the
proposed project site and placed in plastic jars or containers.  If
trace organic constituents are to be determined,  the sample should  be
stored in a glass container or a second sample of approximately the
same size should be collected and stored separately in glass  containers.
Samples should be stored at UC.
            Upon reaching the laboratory, sediment samples may be
stored wet, air dried, or frozen.  The selection between  these
preservation techniques should be based primarily on the  specific

                               2-38

-------
ro
MD
     [SEDIMENT SAMPLE
     INTERSTITIAL WATER
           PHASE
         (Preserve
          Anaerobic
          Integri ty)
Section and
Centri fuge
              -^RESIDUE
      Supernatant
         (Filter)
         Ac i d i fy
         (pH 1-2)

      SOLUTION
      ANALYZE AS
        INTERSTITIAL
        WATER PHASE
                 EXCHANGEABLE  PHASE
EXTRACT with
                             1N  NH,,OAc
                             -Centri fuge
                    (Preserve
                     Anaerobic
                     Integri ty)
                       Washy
RESIDUE
                 Supernatant
                    (Filter)
                    Acidi fy
                 SOLUTION

                 ANALYZE AS
                   EXCHANGEABLE
                   PHASE
          EASILY REDUCIBLE
               PHASE
                                      EXTRACT with
                                         NH2OH-HC1
                                                 Centri fuge
                                                  Wash
RESIDUE
          Supernatant
             (Filter)
          SOLUTION

          ANALYZE AS
            EASILY
            REDUCIBLE
            PHASE
            ORGANIC  +
          SULFIDE  PHASE
                             DIGEST with
                                   H202
                                (pH 2.5)
                                at 95C
                                                                 Extract as
                                                                    Exchange-
                                                                    able
                                                                    (pH 2.5)
                               Wash,
ESI DUE
          Supernatant
             (Filter)
          SOLUTION

          ANALYZE AS
            ORGANIC +
            SULFIDE
            PHASE
           MODERATELY  REDUCIBLE
                    PHASE
                          EXTRACT with
entrifuge
                                                                                            Uoch
         Supernatant
            (Filter)
         SOLUTION

         ANALYZE AS
           MODERATELY
           REDUCIBLE
           PHASE
                                                                          ESIDUE
                                                                                  RESIDUAL  PHASE
                                    DIGEST at 95C
                                       with HF +
                                       HN03 +
                                       (Fuming)
                                       HN03
(Centrifuge)
                                                                                    '
                                                                                   SOLUTION

                                                                                   ANALYZE AS
                                                                                     RESIDUAL
                                                                                     PHASE
                     Figure 2-2.     Elemental partitioning  for sediment characterization

-------
parameter to be determined and, secondarily, on personal choice.
Several parameters such as pH, redox, total solids, and volatile
solids must be run on wet samples.   Other parameters may change due
to oxidation (chlorine demand, BOD, COD, SOD, "sulfides), volatili-
zation (phenolics, volatile solids), or chemical instability (carbamates,
herbicides).  Samples to be analyzed for these parameters should be
processed as soon as possible using subsamples of the original wet
sample.  Samples to be analyzed for particle size (dispersed), total
organic carbon (TOG), metals (except possibly mercury), chlorinated
hydrocarbon pesticides, and PCB's may be stored wet, dried, or frozen.
            Apparatus.   The specific equipment necessary will vary
depending on the chemical constituent(s) to be analyzed in the total
sediment digest or the total sediment organic extract.  Specific
needs and cleanup procedures are discussed with each parameter in
Section 3.
            Test procedure.  The following stepwise procedure is
recommended for the processing of sediment or fill material samples
to be analyzed for total or bulk content:
            a_.  Decant any overlying water that may have been collected
                with the dredge or corer.
            b_.  Blend the dredge, core, or sectioned core sample.
            c_.  Transfer an aliquot of the homogenized sample to a
                tarred weighing dish and weigh.  Dry the sample at
                105C to a constant weight.  This information will
                allow calculation of percent solids in the sample
                and to report subsequent bulk analysis results on a
                milligram-per-kilogram dry weight basis.  The dried
                sample from the percent solids determination may be
                subjected to further chemical analysis for those
                parameters not affected by the drying process.
                If volatile solids are to be determined, record the
                weight of the crucible and the dried sample used in
                the percent solids determination.  Place the sample
                in an electric muffle furnace and ignite the sample
                at 600C for 60 min.  Remove the sample from the
                furnace, allow to cool, and desiccate for 30 min
                prior to weighing.   Report the weight lost on
                ignition as percent volatile solids.
            d_.  Transfer a second subsample of the blended sample to
                a suitable container for pH and oxidation-reduction
                (redox) potential determinations.  The sample size
                                2-1*0

-------
                should be sufficient to allow the electrodes to be
                inserted to a depth of k to 6 cm.  Allow sufficient
                time for the electrode responses to stabilize and
                record the respective pH and redox values.

            e_.  Set aside subsamples of the wet, blended sample for the
                analysis of time-dependent or unstable chemical
                constituents.  Parameters in this category include
                biological oxygen demand, chemical oxygen demand,
                sediment oxygen demand, chlorine demand, herbicides and
                carbamates, phenolics, sulfides, nitrogen,  phosphorus,
                and oil and grease.  Thus, as many as 12 subsamples
                (if all listed analyses are to be performed) will be
                required.  Suggested sample sizes for each aliquot are
                presented in Figure 2-3.  These analyses should be
                initiated as soon as possible to minimize the effects
                of sample alteration due to handling and storage.
            f_.  Set aside separate subsamples for the analysis of
                particle size, carbon, metals, and chlorinated
                hydrocarbons.  However, because of the increased
                stability of these constituents (relative to those
                in Step e_, above), the aliquots may be taken from the
                initial wet, blended sample, or a sample that has been
                dried or frozen for storage.  Required subsample sizes
                are presented in Figure 2-3.
            Individuals performing bulk analysis of sediment samples
should be aware of the fact that analytical results may be affected by
sample handling and storage procedures.  The following special caveats
are maintained here because of the importance of this fact and again
with the appropriate analytical procedure in Section 3:
            a_.  It is preferable to determine Eh and pH values in the
                field as soon as the sample is collected since there is
                no way to stablize these parameters.  If this is not
                possible, these parameters should be determined as
                soon as possible in the laboratory using a wet sample.
                Sample handling should be kept to a minimum to avoid
                sample dehydration or sample oxidation.
            b_.  Percent solids and specific gravity also must be
                determined on a sample of original moisture content.
                The sample should be handled in such a manner to
                minimize water loss and sample dehydration.
            c_.  Cation exchange capacity can be influenced by sample
                drying.  Therefore, it is recommended that this
                parameter be determined on original moisture content
                samples.
            d^  Chlorine demand, biological oxygen demand, chemical
                oxygen demand, and sediment oxygen demand are all

                               2-Ul

-------
ro
fr-
ro
                Dredge or Grab
                    Sample
    Particle Size
       25-50 q
    Mi neralogical
     Compos i tion
       10-25 q
  Total
 Organic
  Carbon
    &
  Total
Inorganic
  Carbon
  1-2  q
    Heavy Metals
       1-10 q
       PCB's &
     Pesticides
      10-100 q
                  Dry Sample
                  for storaqe
                  Freeze Sample
                  for storage
Dry Sample
105C
% Sol ids
5-20 q
i
r

Ignite Residue
600 "C
Volati le Sol ids
                         Note:  All sample sizes given on
                                a dry weight basis.
                                                                           Time-dependent Parameters
                                                                              Process  Immediately
                                                                                  Cation  Exchange Capacity
                                                                                         5-10 g
                                                                                  Chlorine Demand
                                                                                         1-2  g
                                                                                  Biochemical  Oxygen Demand
                                                                                         0.5-5.0 q
Chemical Oxygen Demand
       0.5-5.0 q
                                                                           Sediment Oxygen Demand
                                                                                  10-100 q
Herbicides & Carbamates
       10-100 q
                                                                                  Phenol ics
                                                                                         10-50 q
                                                                                  Sulfides
                                                                                         1-5  q
                                                                                  Specific  Gravity
                                                                                         250  q
                                                                           Nitrogen forms
                                                                                  0.5-10 q
                                                                           Phosphorus forms
                                                                                  0.5-10 g
                                                                                 Oil  &  Grease
                                                                                         5-50 q
                          Figure  2-3.    Sediment sample splitting for bulk analysis

-------
    measures of the reducing capacity of the sample being
    analyzed.  Since sediments are frequently reduced and
    contain elevated concentrations of ferrous iron,
    manganous manganese, and sulfide that can be oxidized
    by atmospheric oxygen, these parameters should be run
    on wet samples.

e_.  Herbicides and carbamates are chemically unstable with
    relatively short half-lives.  Immediate extraction of
    the original sample with methylene chloride reduces
    the possibility of chemically or biologically catalyzed
    decomposition and increases herbicide and carbamate
    stability.

f_.  Phenolic compounds may be lost by volatilization during
    storage.  Therefore, samples to be analyzed for phenols
    should not be dried and storage time should be minimized.
    If immediate analysis is not possible, storage by
    freezing may be acceptable.  Subsequent sample thawing
    should be accomplished at low temperature to reduce
    phenol loss by volatilization.

g_.  Sulfides in the sample may be lost by volatilization
    and oxidation.  It is recommended that sample contact
    with atmospheric oxygen be minimized between sample
    collection and analysis to reduce this effect.  This
    can best be accomplished by excluding air bubbles from
    sample containers and minimizing sample storage time.

h..  Some forms of nitrogen that are expected to occur in
    sediments (nitrites) are unstable in the presence of
    oxygen and can be lost on sample drying.  In addition,
    sample composition may be altered by the uptake or
    loss of volatile ammonia.  Therefore, sample processing
    should begin as soon as possible using a sample of
    original moisture content.
i_.  The distribution of phosphorus forms may be altered by
    changes in other sample constituents.  For example,
    the oxidation of iron in a sample may precipitate
    soluble phosphate.  Therefore, if soluble phosphate
    is to be determined, wet samples should be processed
    as soon as possible.  If total phosphate is the only
    parameter of concern, analysis can be conducted on a
    wet, dried-, or frozen sample.
j_.  The oil and grease content of samples may be reduced
    due to volatilization.  Consequently, sample drying
    prior to analysis is not recommended.

It.  The selection of a storage technique for samples to be
    analyzed for particle size depends on the method of
    analysis.  If apparent particle size is to be run,
    a wet sample should be used.  However, if dispersed
    particle size is to be run, a wet, dried, or frozen
    sample may be used.
                     2-U3

-------
            !_.   Most of the heavy metals are stable and samples
                scheduled for analysis can be stored in a wet, dried,
                or frozen state.   The selection of a storage method
                can affect the distribution of a metal among various
                forms, but the total concentration should be unaffected.
                Two possible exceptions are mercury and selenium,  which
                can be lost by volatilization.  This particularly  is
                true if the samples are dried above 60C.

            m.   Chlorinated hydrocarbon pesticides and PCB's are stable
                and probably unaffected by the method of sample  storage.
                Improved stability can be achieved by immediate
                extraction of the original sample with an organic
                solvent and is suggested, but not essential.

            n_.   The analysis of a sediment or fill material sample for
                specific constituents will require a sample digestion
                or sample extraction technique.  Since the selection of
                a digestion solution or solvent is dependent on  the
                analysis to be performed, this information is presented
                with the specific analytical techniques.
                                Summary


            Dredged material may be subjected to several types of

testing.  This section has provided guidance for conducting elutriate

testing, elemental partitioning, and bulk analysis of sedimentary

samples.  Since each of these procedures measures a different property

of the sample, different storage requirements are required for samples

to be subjected to each testing procedure.  Therefore, this section

has also provided detailed guidance for the handling of sedimentary

samples from the time of collection until the time of analysis.

-------
                              References
 1.   Environmental Protection Agency.   "Draft Interim Analytical
     Methods Manual for the Ocean Disposal Permit Program."
     EPA; Washington,  B.C.  (January 197*0?
 2.   Environmental Effects  Laboratory.   "Ecological Evaluation of
     Proposed Discharge of  Dredged or  Fill Material into Navigable
     Waters.  Interim Guidance for Implementation of Section
     of Public Law 92-500 (Federal Water Pollution Control Act
     Amendments of 1972)."   Miscellaneous Paper D-76-17.  U. S.
     Army Engineer Waterways Experiment Station, CE; Vicksburg,
     Mississippi (1976).
 3.   Environmental Protection Agency.   "NPDES Compliance Sampling
     Manual."  Enforcement  Division, Office of Water Enforcement,
     Compliance Branch, EPA; Washington, D.C.  139 p. (l977)
 U.   American Public Health Association.  Standard Methods for the
     Examination of Water and Wasjbewater Including Bottom Sediments
     and Sludges.   l*rth Edition.   American Public Health Association,
     New York, New York. 1193 p. (1975).
 5.   Sly, P. G.  "Bottom Sediment Sampling."  Proc. 12th Conf. Gr.
     Lakes Res.  pp. 883-898  (1969).
 6.   Howmiller, R. P.   "A Comparison of the Effectiveness of Ekman
     and Ponar Grabs."  Trans. Amer. Fish. Soc. 100:560-564 (1971).
 7.   Wigley, R. L.  "Comparative Efficiencies of Van Veen and Smith-
     Mclntyre Grab Samplers as Revealed by Motion Pictures."  Ecology
     U8:168-169 (1967).
 8.   Hudson, P. L.  "Quantitative Sampling with Three Benthic Dredges."
     Trans. Amer.  Fish. Soc. 99:603-607 (1970).
 9-   Christie, N.  D.  "Relationship Between Sediment Texture, Species
     Richness, and Volume of Sediment  Sampled by a Grab."  Marine
     Biology 30:89-96 (1975).
10.   Brannon, J. M., Plumb  Jr., R. H., and Smith, I.  "Long Term Release
     of Contaminants from Dredged Material."  Technical Report D-78-^9.
     U. S. Army Engineer Waterways Experiment Station, CE, Vicksburg,
     Mississippi.   66 p. (1978).
11.   Brannon, J. M., Engler, R. M., Rose, J. R., Hunt, P. G., and
     Smith, I.  "Selective  Analytical  Partitioning of Sediments to
     Evaluate Potential Mobility of Chemical Constituents During
     Dredging and Disposal  Operations."  Technical Report D-76-7.
     U. S. Army Engineer Waterways Experiment Station, CE; Vicksburg,
     Mississippi.   90 p. (1976).
12.   Jenne, E. A., and Luoma, S.  N.  "Forms of Trace Elements in Soils,
     Sediments, and Associated Waters:  An Overview of Their Determi-
     nation and Biological  Availability."  U. S. Geological Survey;
     Menlo Park, California.  Presentation at Biological Implications

                               2-U5

-------
     of Metals in the Environment.   15th Life Sciences Symposium;
     Hanford, Connecticut.  T^ p.  (September 29 - October 1, 1975).

13.  Chen, K. Y., Gupta, S. K., Sycip, A. Z. , Lu, J.  C. S., Knesevic, M.,
     and Choi, W. W.  "Research Study on the Effect of Dispersion
     Settling, and Resedimentation on Migration of Chemical Constituents
     During Open Water Disposal of Dredged Materials."  Contract Report
     D-76-1.  U. S. Army Engineer Waterways Experiment Station, CE;
     Vicksburg, Mississippi.  221 p. (1976).

1^.  Lee, G. F.  "Chemical Aspects of Bioassay Techniques for
     Establishing Water Quality Criteria."  Water Res. 7:1525-15^6
     (1973).
15.  Environmental Protection Agency.  "Methods for Chemical Analysis
     of Water and Wastes."  National Environmental Research Center,
     EPA; Cincinnati, Ohio.  EPA-625/6-7^-003.  298 p. (197*0.
16.  Sherma, J.  "Manual of Analytical Quality Control for Pesticides
     and Related Compounds in Human and Environmental Samples."
     Contract 68-02-1727.  EPA Health Effects Research Laboratory;
     Research Triangle Park, North Carolina.  EPA-600/1-76-017-
     Unnumbered  (1976).

17.  Environmental Protection Agency.  "Handbook for  Analytical
     Quality Control in Water and Waste-water Laboratories."
     Environmental Monitoring and Support Laboratory,  Environmental
     Research Center; Cincinnati,  Ohio.  Draft Report (1978).
18.  Delfino, J. J.  "Quality Assurance in Water and Wastewater
     Analysis Laboratories."  Water and Sewage Works  12U(7):79-8^
     (1977).
19-  Anon.  "Federal Environmental Monitoring:  Will the Bubble Burst?"
     QA Report.  Environmental Science and Technology 12:126^-1269
     (1978).
20.  Keeley, J. W., and Engler, R.  M.  "Discussion of Regulatory
     Criteria for Ocean Disposal of Dredged Materials:  Elutriate
     Test Rationale and Implementation Guidelines."  Miscellaneous
     Paper 0-7^-1^.  U. S. Army Engineer  Waterways Experiment
     Station, CE; Vicksburg, Mississippi (l97^).
21.  Lee, G. F., Piwoni, M. D., Lopez, J. M., Mariani, G. M.,
     Richardson, J. S.., Homer, D.  H. , and Saleh, F.  "Research Study
     for the Development of Dredged Material Disposal Criteria."
     Technical Report D-75-l^.  U.  S. Army Engineer Waterways
     Experiment Station, CE; Vicksburg, Mississippi (1975).
22.  Plumb, R. H., Jr.  "A Bioassay Dilution Technique to Assess the
     Significance of Dredged Material Disposal."  Miscellaneous Paper
     D-76-6.  U. S. Army Engineer Waterways Experiment Station, CE;
     Vicksburg, Mississippi.  16 p. (1976).
23.  Neff, J. W., Foster, R. S., and Slowey, J. F.  "Availability of
     Sediment-Adsorbed Heavy Metals to Benthos with Particular Emphasis
     on Deposit Feeding Infauna."  Technical Report D-78-U2.  U. S.


                                2-l|6

-------
     Army Engineer Waterways Experiment Station,  CE;  Vicksburg,
     Mississippi.   286 p.  (1978).
2k.   Lee, G.  F.  and Plumb, R.  H.  Jr.   "Literature Review on Research
     Study for the Development of Dredged Material Disposal Criteria."
     Contract Report D-7^-1.  U.  S.  Army Engineer Waterways Experiment
     Station, CE;  Vicksburg, Mississippi.  1^5 p. (197^).
                               2-U7

-------
                    SECTION 3:  ANALYTICAL METHODS

                             Introduction

            This section presents analytical procedures for selected
parameters to be used for the analysis of water, sediments, and sediment
fractions.  The procedures can also be used for analysis of biological
tissue with paper sample preparation.  The two major criteria used to
select the procedures were:
            a_.  The procedures have been shown to be precise and
                accurate..
            b_.  The required equipment is generally available.
            The criterion for a precise and accurate procedure was
that it be considered an acceptable test procedure based on guidelines
established in Section 30Mg) of PL 92-500 or, alternately, the pro-
cedure be accepted as standard in analytical compendiums such as
Standard Methods or an ASTM Water Manual based on critical review and
performance evaluation.  A list of acceptable methods and specific
references is provided in Table 3-1.  This list was abstracted from the
Environmental Protection Agency (EPA) list of approved procedures that
has been previously published in the Federal Register.1
            The criterion for equipment availability is more subjective
and based largely on the capabilities of the intended principal users.
The reason for considering this factor is that smaller laboratories
are usually involved in Section kok permit evaluations and these
laboratories are not generally equipped with the more sophisticated
equipment found in larger research laboratories.  Thus, the listed
procedures require colorimeters, atomic absorption spectrophotometers,
and gas chromatographs rather than inductively coupled plasma arc
techniques, neutron activation analysis, or gas chromatograph/mass
spectrophotometers.  The latter equipment can certainly be used but
their distribution is not considered sufficiently widespread to
warrant inclusion in a general manual at this time.
            Each analytical procedure presented in the remainder of
                                  3-1

-------
this section is accompanied with a flow diagram similar to Figure 3-1
and these diagrams have several uses.  First, there are three options for
storing sediment samples:  wet, dry, or frozen.  When there is reason
to believe that one of these methods is unsuitable, such as drying
sediments to be analyzed for oxygen demand, this portion of the flow
diagram has been deleted.  Second, the diagram reinforces the fact that
only wet sediments should be used for the elutriate test, sediment
fractionation studies, and bioassays.  Third, information has been
tabulated on sample containers, preservatives, storage time, digestion
or extraction solutions, and required sample size.   The user is cautioned
that storage times and preservatives for sediments are not known with
certainty.  Therefore, wet sediments should be processed as soon as
possible and preferably within 1 week.  Also, while drying and freezing
may allow extended storage of sediment samples to be analyzed for some
chemical contaminants, the upper limit for such storage is not known
and the general usefulness of the samples is reduced.   (They should
not be used in bioassays or fractionation studies or analyzed for labile
parameters such as oxygen demand, sulfide, and some organic compounds.)
Finally, a uniform sample designation code has been established through-
out the manual as indicated in Figure 3-1.  Thus, Wl is a total water
sample; S1A is an elutriate sample; and S2 is an air-dried sediment
sample.  This code is used in discussing the analytical preparation of
the samples.
            Procedures are also provided for the digestion or extraction
of sediment samples.   The user is cautioned that efficiency of digestion
or extraction solutions is poorly known in a wide range of sediment
types.   This factor was considered in selecting digestion procedures.
For example, hydrofluoric acid digestion is commonly considered as the
most effective digestion procedure for metals.  However, the use of
this acid requires appropriate hoods and safety equipment.  Since this
equipment was not considered routinely available in the laboratories of
the intended users of this manual, a nitric acid-hydrochloric acid
digestion that has been shown to be effective for metals was presented
for use.  While the nitric acid-hydrochloric acid treatment may produce

                                  3-2

-------

WATER SAMPLE
f
* * *
ACIDIFY FILTER N TR"E
J |
STORE ACIDIFY
1 i
D 1 GE5T STORE **

i i
ANALYZE ANALYZE
(Wl) (W2)

W SAMPLE DESIGNATION Wl
U)
PURPOSE Total Water
Cone.

DREDGE
1
CO

SAMPLE
*
'
"T STORE WET
i
^ i


1 J
r
p ^V
ONATE (sir}
;
ANALYZE ANALYZE
(S1A) (SIB)

W2 W3
RE SAMPLE
|
^^^^^J
CORE SECTION
t
*
DRY
I
STORE
 1 1
~1 1
DIGEST DIGEST
1 i
ANALYZE ANALYZE
(SID) (S2)

S1A SIB SIC SID S2

^^^
FREEZE
1
STORE
, 1
DIGEST
i
ANALYZE
(S3)

S3


Soluble Used In Mobile Chemical Bioavail- Total Total Total
Water Elutriate Cone. Distribution ability Sediment Sediment Sediment
Cone. Cone. Cone. Cone.
CONTAINER
SAMPLE TREATMENT
PRESERVATIVE
STORAGE TIME
DIGESTION SOLUTION
SAMPLE VOLUME OR WEIGHT
                  Figure 3-1.      Schematic  of generalized  sample handling  procedure

-------
lower analytical results than hydrofluoric acid treatment for some
samples, the difference is not considered significant in terms of the
present inability to relate bulk analysis results with environmental
effects.  Thus, the procedure is considered suitable for routine use,
safety, and reproducibility.  Where there is a lack of agreement on
the best sample preparation method, several digestion solutions or
solvents are presented.
            One final point should be mentioned.  The methodology
provides for the preparation of three water samples (total water, W-l;
filtered water, W-2; and elutriate, S1A) and as many as five sediment
samples (fractionation, SIB; bioassay, SIC; wet sediments, SID; dried
sediments, S2; and frozen sediments, S3).  It is not necessary to
analyze each of these fractions for all samples collected.  The
fractions are listed only to provide appropriate guidance on their
preparation after it has been decided to run them.  In addition, the
method section provides guidance for UU parameters.  It is not necessary
or recommended that all possible analyses be run on all samples.  This
manual is simply presenting recommended methodologies that should be
followed once it has been determined that a specific analysis is
required.  Also, failure to list a specific analysis does not mean
that it should not be considered for use where appropriate.  However,
other references will have to be used for detailed guidance.

                              References
1.  Environmental Protection Agency.  "Water Programs. Guidelines
    Establishing Test Procedures for the Analysis of Pollutants."
    Federal Register,  p. 52780-52786 (l December 1976)..

-------
                                               Table  3-1

                                     Acceptable Test Procedures*

Parameter and Units
Ammonia (as N),
mg/1
Method**
Manual distillation7
(at pH 9.5) followed
by nesslerization,

EPA
Methods2 .
159
165
168

lUth Ed.
Standard
Methods 3
UlO
Ul2
616
References
(Page Nos. )
Pt. 31
1975 USGS
ASTM1* Methods5
237 116

Other
Approved
Methodsf
6(6iU)
Biochemical oxygen
 demand, 5-d (BODs),
 mg/1

Chemical oxygen
 demand (COD),  mg/1

Chlorinated organic
 compounds (except
 pesticides), mg/1

Hydrogen ion (pH)
 units
 titration, electrode,
 automated phenolate

Winkler (azide modifi-
 cation) or electrode
 method

Dichromate reflux
Gas chromatography
                  11
Electrometric measure-
 ment
 20
239
550
U60
h!2
178
                                7(50)8    9(17)
        12U
           6(610)
129
                   6(606)
                                              (Continued)
                                                              (Sheet 1  of lU)
 * Information in this table abstracted from the Environmental Protection Agency, 1976  (Table Ref l).
 t Number in parentheses refers  to the page number  of  the  indicated  reference  on Page 12, 13, and lU of
  this table.
** Raised numbers refer to the corresponding footnote  on Page 12, 13, and lU of this table.

-------
                                         Table 3-1 (Continued)
Parameters and Units

Kjeldahl nitrogen
 (as N), mg/1
Arsenictotal,mg/
Arsenicdissolved
Cadiumdissolved,
 mg/


Cadmiumdissolved,
Calciumtotal,
       Method**
Digestion and distilla-
 tion followed by
 nesslerization, titra-
 tion, or electrode;
 automated digestion,
 automated phenolate
Digestion followed
 by silver diethyl
 dithocarbamate or
 atomic absorption13 15
O.H5-y filtration14
 followed by refer-
 enced method for
 total arsenic
Digestion12 followed
 by atomic
 absorption13 >15

O.U5-y filtration14
 followed by refer-
 enced method for
 total cadmium
Digestion12 followed
 by atomic
 absorption13 >15
  EPA
Methods2

  175
  165
  182
                                                    95
  101
  103
                                                                     References
                                                                     (Page Nos.)
lUth Ed.
Standard
Methods 3

  U37
             285
             283
             159
  1U8
  182
  1U8
  189
Pt. 31
 1975
ASTM4
  USGS
Methods 5

  122
 Other
Approved
Methodst
 6(612)
                      (3D10
                      (37)10
            62
             (619)6
              (37)9
 3U5
   66
                                               (Continued)
                                                             (Sheet 2 of lU)
   Number in parentheses refers to the page number of the indicated reference on page 12, 13, and
   U of this table.

-------
                                         Table 3-1 (Continued)
References
(Page Nos . )
Parameters and Units Method**
EPA
Methods2
lUth Ed.
Standard
Methods 3
Pt. 31
1975
ASTM4
USGS
Methods5
Other
Approved
Methodst
Calciumdissolved,
  mg/
Chromium VI,  mg/1
Chromium VI
 dissolved, mg/1


Chromiumtotal,
 mg/1
O.U5-U filtration14 fol-
 lowed by referenced
 method for total
 calcium

Extraction and atomic        89
 absorption; colori-        105
 metric
 (Diphenylcarbazide)

O.U5-y filtration14
 followed by referenced
 method for chromium VI

Digestion12 followed by     105
 atomic absorption13 or
 by colorimetric
 (diphenylcarbazide)
192
           76
           75
1U8
192
286
78
77
                    6(619)
                                             (Continued)
                                                             (Sheet 3 of lU)
t  Number in parentheses refers to the page number of the  indicated reference  on  page 12,  13, and
  lU of this table.

-------
                                              Table 3-1 (Continued)
u>
CO

Parameters and Units
Chromium  dissolved,
mg/1

Copper  total ,
mg/1
Copper  dissolved ,
mg/1
Iron  total,
mg/1
Iron  dissolved,
mg/1
Lead  total ,
mg/1



EPA
Method** Methods 2
0.^5- V filtration1"
followed by referenced
method for total
chromium
Digestion12 followed by 108
atomic absorption13 or
by color ime trie
(neocuproine)
0.1*5-y filtration1"
followed by referenced
method for total copper
Digestion12 followed by 110
atomic absorption1 3
or by colorimetric
( phenanthroline )
OA5-y filtration1"
followed by referenced
method for total iron
Digestion12 followed by 112
atomic absorption13 or
by colorimetric
(dithizone)
(Continued)
References
(Page Nos. )
lUth Ed. Pt. 31 Other
Standard 1975 USGS Approved
Methods3 ASTM" Methods5 Methodst


1U8 3^5 83 6(619)
196 293 9(37)

1U8 3U5 102 6(619)
208 326

1U8 3^5 105 6(619)
215
(Sheet h of U)
         Number in parentheses refers to the page number of the indicated reference on Page 12,  13,  and Ih  of
        this table.

-------
                                        Table 3-1 (Continued)
Parameters and Units

Leaddissolved,
 mg/1
Magnesiumtotal,
 mg/1
Magnes iumdi s solved,
 mg/1
Manganesetotal,
 mg/1
Manganesedissolved,
 mg/1
                                                                    References
                                                                    (Page Mos.)
Method**
   EPA f
Methods'
lUth Ed.
Standard
Methods 3
Pt. 31
 1975
ASTM4
                                                         USGS
                                                       Methods'
  Other
Approved
Methodst
O.U5-y filtration11*
 followed by referenced
 method for total lead.
Digestion12 followed by    llU
 atomic absorption; or
 gravimetric

O.U5-y filtration11*
 followed by referenced
 method for total
 magnesium
Digestion12 followed by    116
 atomic absorption13 or
 by colorimetric
 (persulfate or
 p.eriodate)

O.U5-y filtration11*
 followed by referenced
 method for total
 manganese

                     (Continued)
                                1U8
                                221
                                  100
                                6(619)
                                1U8
                                225
                                227
                       31*5
                       111
                     6(619)
                                                                                   (Sheet 5 of 1U)
t  Number in parentheses refers to the page number of the indicated reference on Page 12, 13, and 1U of
  this table.

-------
                                              Table 3-1  (Continued)
U)

M
O
References
(Page Wos . )
Parameters and Units
EPA
Method** Methods2
iVth Ed.
Standard
Methods 3
Pt. 31
1975
ASTM1*
USGS
Methods5
Other
Approved
Methodst
       Mercurytotal, mg/1
       Mercurydi s solved,
        mg/1
Molybdenumtotal,
 mg/1

Molybdenumdi s solved,
 mg/1
       Nickeltotal,
        mg/1
       Ni ckel di s s olved,
        mg/1
 156
Flameless atomic          118
 absorption

O.U5-y filtration14
 followed by referenced
 method for total
 mercury

Digestion12 followed by   139
 atomic absorption13

O.U5-y filtration11*
 followed by referenced
 method for total
 molybdenum

Digestion12 followed by   1^1
 atomic absorption13 or
 by colorimetric
 (heptoxime)

O.U5-y filtration16
 followed by referenced              232
 method for total
 nickel
                     (Continued)
lU8
                                                                       338
11(5D10
                                                                              350
                                                                       3U5
   115
                                                                                           (Sheet 6 of- lU)
          Number in parentheses refers  to the page number of the  indicated  reference on Page 12, 13, and 1U  of
         this table.

-------
                                        Table  3-1  (Continued)
Parameters and Units

Potassiumtotal,
 mg/1
Potassiumdissolved,
 mg/1
Seleniumtotal,
 mg/1
Seleniumdi s solved,
 mg/1
Sodiumtotal, mg/1
                               Method**
                                                                    References
                                                                    (Page  Nos. )
Digestion12 followed
 by atomic absorption,
 colorimetric
 (cobaltinltrite), or
 by flame photometric
0.^5-y filtration11*
 followed by referenced
 method for total
 potassium
Digestion12 followed by
 atomic absorption15'16
0.1*5-y filtration1"
 followed by referenced
 method for total
 selenium

Digestion12 followed by
 atomic absorption or
 by flame photometric
   EPA
Methods2

  1U3
                                                               Ed.
                                                          Standard
                                                          Methods 3
                                                              235
                                                  1U5
159
                                                              250
Pt. 31
 1975
ASTM**

 U03
                   USGS
                 Methods
  Other
Approved
Methodst

 6(620)
        U03
                                                                                1U3
                                          6(621)
                                             (Continued)
                                                                                    (Sheet 7  of 1U)
t  Number in parentheses refers  to  the  page  number  of the  indicated reference  on Page 12, 13, and
  this table.
                                                                                                     of

-------
                 Table 3-1 (Continued)
Parameter and Units

Sodiumdissolved,
 mg/1
Zinctotal,  mg/1
Zinc-dissolved,
 mg/1


Nitrate (as N),
 mg/1
Nitrite (as N),
 mg/1
                                             Reference
                                             (Page Nos . )
        Method**
                                                 EPA
                                               Methods2
lUth Ed.
Standard
Methods3
                                                                      Pt.  31
                                                                       1975
                                                                      &STM11
                                                                                 USGS
                                                                               Methods
                                                                                            Other
                                                                                          Approved
                                                                                          Methodsi
0.1*5-u filtration1"1
 followed by referenced
 method for total
 sodium

Digestion12 followed by    155
 atomic absorption13 or
 by colorimetric
 (dithizone)

O.U5-y filtration11*
 followed by referenced
 method for total zinc

Cadmium reduction;
 bruc i ne sulfat e;
 automated cadmium
 or hydrazine
 reduction
                                       lU8
                                       265
                                                                       3^5
                       159
                                                                                           6(619)
                                                                                           9(37)
                           201
                           197
                           207
                                                              U23
                                                              U27
                                                              620
            358
                                                                                  119
                                                                                           6(6lU)
                                                                                           9(28)
          17
Manual or automated
 colorimetric
 (diazotization)
                                                  215
                      (Continued)
                       121
                                                                                    (Sheet 8  of  1U)
t  Number in parentheses refers  to  the  page  number  of  the indicated reference on Page 12, 13, and
  this table.
                                                                              of

-------
                                             Table  3-1  (Continued)
u>
References
(Page Nos. )

Parameters and Units
Oil and grease, mg/1



Organic carbon (as
TOG), mg/1
Organic nitrogen
(as N) , mg/1

Ort ho -p ho sphat e
(as P), mg/1

Pentachlorophenol ,
mg/1
Pesticides, mg/1
Phenols , mg/1


Method**
Liquid-liquid
extraction with
trichchlorotrifluoro-
ethane-gr,avimetric
Combustion infrared
method l 8
Kjeldahl nitrogen
minus ammonia
nitrogen
Manual or automated
ascorbic acid
reduction
Gas chromatography 1 l
Gas chromatography 1 l
Colorimetric (ItAAP)


EPA
Methods 2
229



236
175
179

2U9
256



2Ul
(Continued)
lUth Ed. Pt. 31
Standard 1975
Methods 3 ASTM1*
515



532 U67
1*37

U8l 384
62U


555 529
57^ 5^5

Other
USGS Approved
Methods5 Methods t




17(10
122 6(612)
6(6lU)

131 6(621)


19(2U)t

(Sheet 9 of 1U)
     t  Number in parentheses refers to the page number  of  the indicated reference on Pagel2,  13, and lU  of
       this table.

-------
                                              Table 3-1 (Continued)
U)
J=-
References
(Page Nos. )
Parameters and Units
Pho sphorus  total
(as P), mg/1
Solids  total ,
mg/1
Solids  total dissolved
(filterable), mg/1
Solids  total sus-
Method**
Persulfate digestion
followed by manual
or automated ascorbic
acid reduction
Gravimetric, 103 to
105C
Glass fiber filtration,
180C
Glass fiber filtration,
EPA
Methods 2
256
270
266
268
itth Ed. Pt. 31
Standard 1975
Methods 3 ASTM4
1+76 38U
1*81
62U
91
92
9U
Other
USGS Approved
Methods5 Methods f
133 6(621)



       pended (non-
       filterable), mg/1

      Solidssettleable,
       ml/1 or mg/1

      Solidstotal
       volatile, mg/1

      Specific conductance,
       umhos/cm at 25C
 103 to 105C


Volumetric or gravi-
 metric
Gravimetric, 550C
Wheatstone bridge
 conductimetry
272


275
             95
95
71
120
1U8
6(606)
                                                   (Continued)
                                                            (Sheet 10 of lU)
      t  Number in parentheses refers to the page number of the indicated reference on Page.12, 13, and lit of
        this table.

-------
                                              Table 3-1 (Continued)
                                                                          References
                                                                          (Page Nos. )
                                                                 lUth Ed.   Pt. 31                Other
                                                         EPA     Standard    1975      USGS     Approved
      Parameters and Units           Method**         Methods2   Methods3   ASTM"    Methods5   Methodst
       Sulfide  (as S),        Titrimetriciodine for     2&k        505
       mg/1                   levels greater than                   503
                              1 mg/1; methylene-
                              blue photometric

       Temperature, C        Calibrated glass or         286        125               20(3l)t
                              electrometric
                              thermometer
uo
H
v/i
                                                    (Continued)                           (Sheet 11 of 1U)
           Number  in parentheses refers to the page number of the indicated reference on Page 12, 13, and
          1U of this table.

-------
                                            Table 3-1 (Continued)
                                            References

 1.   Environmental Protection Agency.   "Water Programs Guidelines Establishing Test Procedures for
     the Analysis of Pollutants."  Federal Register, 52780-52786  (l December 1976).

 2.   Environmental Protection Agency.   "Methods for Chemical Analysis of Water and Wastes."
     Environmental Monitoring and  Support Laboratory, Office of Research and Development, EPA;
     Cincinnati,  Ohio.   298 p.  (197U).

 3.   American Public Health Association.  Standard Methods for the Examination of Water and
     Wastewater Including  Bottom Sediments and Sludges.  lUth Edition.  American Public Health
     Association, New York, New York.   1193 p. (1975).

 U.   American Society for  Testing  and Materials.  Annual Book of Standards, Part 31, Water.
     American Society for  Testing  and Materials, 19l6 Race Street, Philadelphia, Pennsylvania  (1976).

 5.   Brown,  E., Skougstad, M. W. ,  and Fishman, M. J.  "Methods for Collection and Analyses of Water
     Samples for Dissolved Minerals  and Gases."  U. S. Geological Survey Techniques of Water
     Resources Inventory,  Book  5,  Chapter Al.  Reston, Virginia (1970).  All page citations refer
     to this reference unless otherwise noted.

 6.   EPA comparable method may  be  found on the indicated page of "Official Methods of Analysis of
     the Association of Official Analytical Chemists," methods manual, 12th ed. (1975).

 7.   Manual distillation is not required if comparability data on representative effluent samples
     are on file to show that this preliminary distillation step is not necessary; however, manual
     distillation will be  required to resolve any controversies.

 8.   Slack,  K. V. et al.   "Methods for  Collection and Analysis of Aquatic Biological and Micro-
     biological Samples."  U. S. Geological Survey Techniques of Water Resources Inventory, Book 5,
     Chapter AU.   Reston,  Virginia (1973).

 9.   American National Standard on Photographic Processing Effluents, April 2, 1975.  Available from
     MSI, 1U30 Broadway;  New York,  New York 10018.

10.   Fishman, M.  J., and Brown, E.  "Selected Methods of the U. S. Geological Survey for Analysis

                                            (Continued)                            (Sheet 12 of lU) .

-------
                                       Table  3-1  (Continued)
                                      References  (Continued)

     of Wastewaters".   Open-file  report  76-177  (1976).
11.  Procedures for pentachlorophenol, chlorinated  organic compounds, and pesticides can be obtained
     from the Environmental Monitoring and Support  Laboratory, EPA;  Cincinnati, Ohio U5268.

12.  For the determination of total metals, the sample  is not filtered before processing.
     Because vigorous  digestion procedures may result in a loss of certain metals through
     precipitation, a  less vigorous treatment is  recommended as given on p. 83  (U.l.U)  of
     "Methods for Chemical Analysis of Water and  Wastes" (197*0 .  In those instances where
     a more vigorous digestion is desired, the procedure on p. 82 (U.I.3) should be followed.
     For the measurement of the noble  metal series  (gold, iridium, osmium, palladium, platinum,
     rhodium, and ruthenium), an  aqua  regia digestion is to be substituted as follows:  Transfer
     a representative  aliquot of  the well-mixed sample  to a Griffin  beaker and  add 3 ml of
     concentrated redistilled HN03. Place the beaker on a steam bath and evaporate to  .dryness.
     Cool the beaker and cautiously add  a 5--ml portion  of aqua regia.  (Aqua regia is prepared
     immediately before use by carefully adding three volumes of concentrated HC1 to one volume
     of concentrated HN03.)  Cover the beaker with  a watch glass and return to  the steam bath.
     Continue heating  the covered beaker for 50 min.  Remove cover and evaporate to dryness.
     Cool and take up  the residue in a small quantity of 1:1 HC1.  Wash down the beaker walls
     and watch glass with distilled water and filter the sample to remove silicates and other
     insoluble material that could clog  the atomizer.   Adjust the volume to some predetermined
     value based on the expected  metal concentration.   The same is now ready for analysis.

13.  As the various furnace devices (flameless automic  absorption spectrophotometer) are essentially
     atomic absorption techniques, they  are considered  to be approved test methods.  Methods of
     standard addition are to be  followed as noted  on p. 78 of "Methods for Chemical Analysis  of
     Water and Wastes" (197*0.

lU.  Dissolved metals  are defined as those constituents that will pass through  a O.U5-y membrane
     filter.  A Prefiltration is  permissible to free the sample from larger suspended solids.
     Filter the sample as soon as practical after collection using the first 50 to 100  ml to rinse

                                            (Continued)                           (Sheet 13 of

-------
                                      Table 3-1  (Concluded)
                                      References  (Continued)

     the flask and collect  the  required volume of  filtrate.  Acidify the  filtrate  with 1:1
     redistilled HNC-3  to a  pH of  2.  Normally, 3 ml of  (l:l) acid per  liter  should be sufficient
     to preserve the samples.

15.  See "Atomic Absorption Newsletter," vol. 13,  75  (19?U).  Available from Perkin-Elmer
     Corporation, Main Avenue,  Norwalk, Connecticut 06852.

16.  Method available from  Environmental Monitoring and Support Laboratory,  EPA; Cincinnati,
     Ohio 1*5268.

IT.  An automated hydrazine reduction method is available from the Environmental Monitoring and
     Support Laboratory, EPA, Cincinnati,  Ohio U5268.

18.  A number of such systems manufactured by various companies are considered to  be  comparable
     in their performance.   In  addition, another technique, based on combustion-methane
     detection, is also acceptable.

19-  Goerlitz, D., and Brown, E.   "Methods for Analysis of Organic Substances in Water."
     U. S. Geological Survey Techniques of Water Resources Inventory,  Book 5, Chapter A3  (1972).

20.  Stevens, H. H., Ficke, J.  F.,  and Smoot, G. F.   "Water TemperatureInfluential  Factors,
     Field Measurement and  Data Presentation".  U. S. Geological Survey Techniques of Water
     Resources Inventory, Book  1  (1975).
                                                                                  (Sheet 1U of lU)

-------
PHYSICAL ANALYSIS

     Cation Exchange Capacity

     Particle Size

     pH

     Oxidation Reduction Potential

     Solids
          Total
          Volatile

     Specific Gravity
                                 3-19

-------
                       CATION EXCHANGE CAPACITY

            The cation exchange capacity (CEC) of a sediment is a measure
of the reversibly bound cations in the sample, that is,  a measure of
those cations held on the surface, within the crystaline matrix of some
minerals.l*  These cations may potentially be released to the vater
column under appropriate conditions.
            The procedure consists of equilibrating a sediment sample
with a highly soluble salt solution.1"3  The theory behind the procedure
is that the high concentration of a soluble cation will  replace the
sorbed or bound cations associated with the sediment. The replaced
cations can then be determined individually in the leachate or the
sediment sample can be washed and reequilibrated with a  second soluble
salt.  The second leachate is then analyzed for total cation exchange
capacity.  The standard leachate that is most often used is 1 N_ ammonium
acetate.  Principal advantages of this approach are the  pH buffering
capacity of ammonium acetate solutions and the relative  ease of the
ammonia determination.  However, ammonium acetate may yield low results
with (a) samples containing 1:1 type clay minerals such  as kaolin or
halloysite, or (b) highly calcareous sediments due to the dissolution
of calcium carbonate.*
            The exchange capacity of a sample is influenced by the clay
content of the sample, the type of clay, the organic matter content,
the pH of the displacing solution, the nature and concentration of the
displacing cation, and the sediment-to-solution ratio.   Since many of
these factors are operationally defined, the CEC of the  sample should
be considered operationally defined.  Most techniques will only vary
the magnitude of the CEC of a sample and not the relative order of a
number of samples.l  Therefore, care should be taken to  standardize as
many variables as possible (ammonia concentration, pH, solid-liquid
ratio, and time of contact) to ensure uniformity and comparibility of
results.

* References for this procedure are on Page 3-27-

                                 3-20

-------
                      Sample Handling and Storage

            Samples may be collected with any convenient collection
device and stored in either glass or plastic containers.  Field moist
samples should be used for the CEC determination as the process of
drying has been shown to alter the CEC.1 **  At this time, the storage
time limits are not known.  Since sample oxidation may indirectly
affect CEC, it is recommended that samples be processed as soon as
practical.  Samples may be frozen, if necessary, but should be thawed
quickly prior to analysis, and portions of samples indicating oxidation
should be discarded.  This information is summarized in Figure 3-2.
                                3-21

-------
CORE SAMPLE

WATER SAMPLE DREDGE SAMPLED
4, 1 1

ACIDIFY STORE WET
1
STORE
1
1
i
CORE S
^
ECTIONJ
1

1 -
DIGEST DIGEST
1 J

ANALYZE ANALYZE
(W1) (SID)

^j SAMPLE DESIGNATION W1 W2 W3 S1A SIB SIC
r PURPOSE <'' Total Water
Cone.
SID
Total
Sediment
Cone.

FREEZE
1
STORE
1
DIGEST
1
ANALYZE
(S3)

S2 S3
Total
Sediment
Cone.
CONTAINER
SAMPLE TREATMENT
PRESERVATIVE
STORAGE TIME
DIGESTION SOLUTION
SAMPLE VOLUME OR WEIGHT
                          G,P
                          None
                                                                                            G,P
                                                                                            None
                         NH OAc
                                                                                          NH OAc
                                                                                          5-25 g
  G,P
                                                                                                                Freeze
                                                                                                                 None
                                                                                                              NH,. OAc
5-25 9
      Figure  3-2.      Handling  and  storage procedures for cation  exchange capacity samples

-------
               Procedures for Sediment Samples (SID, S3)

Method 1:  Agitation, Filtration
Apparatus
Wrist-action shaker
Filtration apparatus
Erlenmeyer flasks
Reagents
1 N_ ammonium acetate:  dissolve 77 g ammonium acetate in distilled water
      and dilute to 1 .
80 percent ethanol:  mix 80 ml ethanol with 200 ml distilled water.
10 percent sodium chloride:  dissolve 100 g sodium chloride in distilled
      water and dilute to 1 I.
Procedure
            Blend sediment sample and pass through a 100-mesh stainless
steel screen to remove coarse particles such as wood and shellfish
fragments.  As quickly as possible (to minimize sample oxidation
effects), weigh out 5- to 25-g subsamples.  The smaller size is suitable
for high silt-clay content sediments and the larger size is suitable
for sandy sediments.
NOTE:  If marine or brackish sediments are used, wash the sediments with
       distilled water until there is no trace of chloride.  Proceed
       as indicated below.
            Rapidly transfer the sample to a 250- to 300-ml Erlenmeyer
flask and immediately add 100 ml 1 N_ ammonium acetate.  Seal the samples
with parafilm and place on a wrist-action shaker.  Agitate the samples
for 30 min.
            While the samples are on the shaker, weigh out a separate
portion of the original sample for a percent solids determination.
Dry the sample at 105C, cool in a desiccator, and reweigh (Page 3-58).
            After 30 min, filter the ammonium acetate-sediment
suspensions through a Whatman No. kO filter or equivalent.  Wash the
retained solids with an additional UOO ml 1 IT ammonium acetate.  Add
the rinsing to the filtrate and retain for the determination of
exchangeable metals if desired.
                                3-23

-------
            Wash the solids vith 25 ml 80 percent ethanol and discard
the filtrate.  Repeat the washing procedure with four additional 25-ml
aliquots of ethanol to ensure complete removal of any excess ammonium
acetate.
            Leach the solids on the filter with 10 percent sodium
chloride solution until 50 ml of leachate has been collected.  Filter the
sample through a 0.^5-M pore-size membrane filter and analyze the
leachate for ammonia using either one of the procedures presented else-
where or an ammonia electrode.
            If it is desired to determine the exchangeable amounts of
specific metals, transfer the original ammonium acetate filtrate to an
appropriate-sized beaker.  Rinse the collection flask with 10 ml
10 percent acetic acid and add to the beaker.  Evaporate the samples to
dryness on a steam bath to remove the ammonium acetate.
            Digest the residue in 10 ml concentrated HWOs and 3 ml
concentrated HClOi*.   When white HClOt* fumes are evolved, remove the
samples and allow to cool.
            Add a small amount of distilled water to the digestate and
filter through a O.h^-li pore-size membrane filter.  Collect the
filtrate in a volumetric flask.  Rinse the beaker with distilled water,
filter, and add to the volumetric flask.
            Dilute to volume with distilled water and analyze for the
metal(s) of choice.
Calculations
            The CEC  of the sediment is reported in meq/100 g and is
calculated as follows:
                           =  (X ag/A) (0.5) (100)
                              (18 mg/rneq) (g) (% S)
where
  X = ammonia concentration in NaCl leachate, mg/C
0.5 = volume of NaCl leachate, B
 18 = millequivalent weight of ammonium ion, mg/meq.
  g = weight of sediment sample,  g
% S = percent solids in sediment  sample (as decimal fraction)

-------
            The exchangeable metal concentration (EMC) is calculated as
follows :
where
  y = metal concentration in the ammonium acetate leachate, mg/.d
  v = final volume of acid digest, H
meq. = milliequivalent weight of metal , mg/meq.
  g = weight of sediment sample, g
% S = percent solids in sediment sample (as decimal fraction)

Method 2:  Centrifugation
            The following method for CEC determination is essentially
the same as the first method except phase separations are accomplished
with a centrifuge rather than filtration.  Reagents are the same.
Procedure
            Weigh out a 5-g sample of homogenized sediment and transfer
to a 50-ml centrifuge tube.
            Add 33 ml 1 N_ ammonium acetate solution,  (it has been
found to be convenient to use a repipet for this procedure.)  Shake
each sample and let the suspension stand for 30 min.  Shake the sus-
pensions and centrifuge for 10 min at 2000 rpm with a table top centri-
fuge.  Decant the ammonium acetate solution and save for exchangeable
metal concentrations.
            Repeat the above procedure with a second and a third 33-ml
portion of ammonium acetate.  Combine the ammonium acetate solutions.
            Add 33 ml 80 percent ethanol to the sediment residue in the
centrifuge tube.  Shake the tubes and centrifuge for 10 min at 2000 rpm.
Decant the ethanol layer and discard.  Repeat the ethanol washing
procedure two times.
            Add 33 ml 10 percent sodium chloride solution to the
washed sediment residue and shake.  Centrifuge the sample and decant
the liquid phase into a 100-ml volumetric flask.  Repeat the process
with two 33-ml portions of 10 percent sodium chloride.  Add the sodium
                                 3-25

-------
chloride decantate to the volumetric flask and dilute to volume.
            Analyze the sediment leachate for ammonia.
            If specific exchangeable metals are to be determined,
evaporate the combined ammonium, acetate extract to dryness on a steam
bath.  Add 100 ml concentrated HNOa and 3 ml concentrated HClOi,.  Heat
on a hot plate until HClOi, fumes begin evolving.
            Cool the sample and add a small amount of distilled water.
Filter through a O.U5-y pore-size membrane filter and collect in a
volumetric flask.  Dilute to volume and analyze for the metal(s) of
interest.
Calculations
            The calculations are the same as for the first method
except the volume of NaCl leachate is 0.1  instead of 0.5 &.
                                3-26

-------
                          References
Black, C. A.  Methods of Soil Analysis.  American Society of
Agronomy and American Society of Testing Materials; Madison,
Wisconsin.  1572 p. (1965).

Toth, S. J., and Ott, A. N.  "Characterization of Bottom Sediments:
Cation Exchange Capacity and Exchangeable Cation Status."  Envir.
Science and Tech. *i:935-939 (1970).

Jackson, M. L.  Soil Chemical Analysis.  Prentice-Hall, Inc.;
Englewood, New Jersey.  1*98 p. (i960).
Plumb, R. H., Jr.  "A Study of the  Potential Effects of the
Discharge of Taconite Tailings on Water Quality in Lake Superior."
Ph.D. Thesis, University of Wisconsin-Madison.  550 p.  (1973).
                           3-27

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                             PARTICLE SIZE
            Particle-size distribution is a cumulative frequency dis-

tribution or a frequency distribution of relative amounts of particles
                                         i*
in a sample within specified size ranges.    The size of a discrete

particle is usually characterized as a linear dimension and designated

as a diameter.  It should be recognized that the use of sieves and

settling tubes will result in a separation based on particle shape as

well as particle size.  Therefore, the following definitions are

presented for comparison of terms that may appear in the technical
literature:2
            a_.  The nominal diameter of a particle is the diameter of a
                sphere that has the same volume as the particle.

            b_.  The sieve diameter of a particle is the diameter  of a
                sphere equal to the length of the side of a square
                sieve opening through which the given particle will
                just pass.

            c_.  The standard fall velocity of a particle is the average
                rate of fall that the particle would attain if falling
                alone in quiescent, distilled water of infinite extent
                and at a temperature of 2hC.
            d_.  The standard fall diameter, or simply fall diameter, of
                a particle is the diameter of a sphere that has a
                specific gravity of 2.65 and has the same standard fall
                velocity as the particle.

            e_.  The sedimentation diameter of a particle is the diameter
                of a sphere that has the same specific gravity and
                terminal uniform settling velocity as the given particle
                in the same sedimentation fluid.
            f_.  The standard sedimentation diameter of a particle is the
                diameter of a sphere that has the same specific gravity
                and has the same standard fall velocity as the given
                particle.
            g_.  The size distribution, or simple distribution, when
                applied in relation to any of the size concepts,  is
                the distribution of material by percentages or propor-
                tions by weight.
            Particle size may be reported as class, millimeters,  micro-
* References for this procedure are on page 3-^7.


                               3-28

-------
meters, or a phi value.  A comparison of these four size scales is
presented in Table 3-2.  A comparison of instrument capabilities based
on cost and particle size range is presented in Table 3-3.
            The size distribution of sediments can be of importance
because it can affect the distribution of chemicals in the  aquatic
environment.  Specifically, sediments can remove chemical contaminants
from water by the process of sorption.   Further, since sorption is a
surface phenomena, the smaller particle sizes generally have a higher
concentration of these chemical contaminants on a weight/weight basis.
            There is a certain amount of arbitrariness associated with
particle-size analysis.  One method relies on the treatment of the
sample with hydrogen peroxide to destroy organic matter that may be
causing the sediment particles to aggregate.  While this approach will
define the true particle-size distribution of the sample, the results
will not be representative of the surface area potentially  available
for sorption or exchange reactions.  On the other hand, sizing of
sediments without peroxide treatment would yield results more repre-
sentative of the exposed surface area but the apparent particle-size
distribution may be affected by the method of sample handling prior to
sizing.

                      Sample Handling and Storage

            Samples scheduled for particle-size analysis may be stored
in either plastic or glass containers.   The samples should  be chilled
at k to 5C but never frozen prior to analysis.  If samples cannot
be analyzed within a few hours, Lugols solution should be added as a
preservative to minimize the effects of bacterial growth.
            Particle-size analysis of suspended solids in water will
require 500 to 2000 ml.  The exact volume will depend on the suspended
solids concentration of the sample.  The required amount of sediment
will range from approximately 3 to 25 g, depending on the size distri-
bution.  Should the sample contain a large percentage of coarse sand and
gravel, a larger sample size should be used to ensure that  the smaller
                               3-29

-------
                               Table 3-2
       Comparison of Scales Used to Report Particle Size  Results

Class Name
Boulders
Cobbles
Gravel
Very coarse sand
Coarse sand
Medium sand
Fine sand
Very fine sand
Coarse silt
Medium silt
Fine silt
Very fine silt
Coarse clay
Medium clay
Fine clay
Very fine clay
Colloids
Millimeters
Micrometers
Phi Value
>256 <-8
256
64
2.0
1.0
0.50
0.25
0.125
0.062
0.031
0.016
0.008
0.004
0.0020
0.0010
0.0005
<0
- 64
- 2
- 1.0
- 0.50
- 0.25
- 0.125
- 0.062
- 0.031
- 0.016
- 0.008
- 0.004
- 0.0020
- 0.0010
- 0.0085
- 0.00024
.00024


2,000
1,000
500
250
125
62
31
16
8
4
2
1
0.5
<0


- 1,000
- 500
- 250
- 125
- 62
- 31
- 16
- 8
- 4
_ 2
- 1
- 0.5
- 0.24
.24
-8 to -6
-6 to -1
-1 to 0
0 to +1
+1 to +2
+2 to +3
+3 to +4
+4 to +5
+5 to +6
+6 to +7
+7 to +8
+8 to +9
+9 to +10
+10 to +11
+11 to +12
>+12
                               Table  3-3
     Comparison of Particle-Size  Distribution Analytical Methods3
                                                         Approximate
Initial Cost, $   Analytical Method   Size Range, ytm   Analysis Cost, $
   500 -   1,000  Optical microscope
                  Sieves
                  Simple sedimen-
                   tation

     10,000       Optical microscope
                  Electron micro-
                   scope
                  Centrifugal
                   sedimentation
                  Sedimentation

                  Electron micro-
10,000 - 100,000   scope
                  Scanning electron
                   microscope
                  Stream counting
                  Instrumented
                   microscope

                  Scanning electron
    100,000+       microscope

                               3-30
  1 to 1,000
  1 to 1,000

  5 to 1,000

0.5 to 1,000

0.2 to 10

0.2 to 50
0.2 to 50

0.1 to 10

0.1 to 100
0.1 to 100

0.5 to 1,000

0.1 to 100
 20

 40

 40

 20

 20
 20


 40

200
 10

 20


200

-------
size classes are being representatively sampled.
            It is recommended that particle-size  samples not be frozen
or dried prior to analysis (Figure 3-3).   The basis of this recommenda-
tion is that the freezing-thawing cycle or sample drying may cause an
irreversible change in the particle-size  distribution due to oxidation
and/or agglomeration.

                           Method Selection
            Particle-size analysis of a sediment sample will usually
require the use of two or more methods because of the wide range of
particle sizes encountered.  The useful size range and amount of
sample required for each method are presented below:1
 Method     Size Range, mm  Concentration, mg/C  Sediment quantity, g
Sieves       0.062-32
Particle
 Counters   0.0002 - 0.062
V. A. Tube    .062-2
Pipette      0.002 - 0.062
2,000 - 5,000
0.05 - 15-0
 1.0 - 5.0
                               3-31

-------
1
CORE SAMPLE
., f j,

WATER SAMPLE bftEOGE SAMPLE CORE SECTION
1 * *
1 T *
SIEVE STORE WET
1

^
SIEVE
1
ANALYZE ANALYZE
(Wl) (SID)
U)
l^> SAMPLE DESIGNATION Wl SID
PURPOSE Total Water Sediment Particle-
Cone. Size Distribution
CONTAINER
SAMPLE TREATMENT
PRESERVATIVE
STORAGE TIME
SAMPLE VOLUME OR WEIGHT
                          G.P
                          None
                          LugoIs
                        Storage Time Not Critical
                          Unless Bacterial
                           Growth Occurs.
                         0.5-2.0 mi
                                                                                               G.P
                                                                                              None
                                                                                              None
                                                                                              25 g
             Figure 3-3.     Handling  and  storage of  samples for particle size analysis

-------
                 Procedures for Sediment Samples (SID)

Method 1:  Sieving and Electronic Particle Counters
Apparatus
Nest of U. S. standard sieves ranging from #230 (62 y) up to #18 (1000 y)
Ro-Tap apparatus for sieving
Coulter counter or equivalent electronic particle counter equipped with
      a 200- and 15-y aperture
Magnetic stirrer
10-ml beakers
250-ml Erlenmeyer flasks
Evaporating dishes
Balance
Reagents
Calgon solution:  dissolve 50 g commercially available Calgon in 1 H
      distilled water.  Filter solution through a 0.2 y pore-size
      membrane filter prior to use.
NaCl electrolyte solution:  dissolve 25 g NaCl electrolyte .solution in
      1  distilled water.  Filter solution through a 0.2-y pore-size
      membrane filter prior to use.
Procedure
            Blend the sediment sample and weigh out 5-0 to 25.0 g
wet sediment.  Add 5 ml Calgon solution and blend for 30 sec.  Wash
container sides with distilled water and blend for an additional 30 sec.
            Wet sieve the suspension through a #230 U. S. standard
sieve.  Collect the material retained on the sieve for size analysis
(Fraction l).  Collect the filtrate in tared beakers (Fraction 2).
            Dry, cool, and weigh the Fraction 2 samples.  This will
provide information to calculate the weight percent of total silt and
clay in the sample.
            Dry and cool Fraction 1.  Record the weight of this fraction.
Place sample on a nest of standard sieves ranging from a #18 standard
sieve (1000 y) to a #120 standard sieve (125 y).  Weigh each individual
fraction.  This information will allow calculation of the -1 phi-size to
+3 phi-size fractions.
                               3-33

-------
            Weigh out a second 5.0- to 25-g blended aliquot of the
original sample.  Add 5.0 ml Calgon solution and blend as before.   Wet
sieve the sample through a #230 U. S. standard sieve and collect the
filtrate in a 250-ml flask and dilute to volume with distilled water
(Fraction 3).
            Determine the size distribution of Fraction 3 using an
electronic particle counter such as the Coulter counter.  Follow
manufacturer's directions to set up the instrument.   Allow 15 min
for warmup.
            Filter a supply of electrolyte through a 0.2-y filter.  Set
the calibration potential and milliamp controls according to the manu-
facturer's instructions.  Count the filtered electrolyte using a 30-y
tube.  The background count should be less than ^00 with the shield
door closed.
            Place the 200-U tube on the sample stand and turn aperture
slightly clockwise.  Position the tube slightly away from the beaker.
Adjust the calibration potential and milliamp settings, if necessary.
Set the gain switch to auto, 'active channel switch to 15-2, sampling
switch to time, mode switch to volume, and display gain switch to X10.
Set the stirring motor to a proper rpm and be sure the propeller is
centered in the bottom of the beaker.  (The mixing process should not
cause surface turbulence.)
            Place Fraction 3 on a magnetic stirrer and mix.  While the
sample is being mixed, withdraw three equal-sized subsamples from the
suspension:  one from the top, middle, and bottom of the flask.   Never
take the bottom subsample against the bottom of the flask.  The sub-
sample volume will depend on the sediment concentration but 3- to 5-ml
aliquots should provide adequate sample.
            Transfer the subsamples to a clean 200-ml beaker and dilute
to volume with filtered 2.5 percent NaCl electrolyte solution.  Using
a 200-y aperture, run the sample through channels 6 to 15 of a Coulter
counter for 60 sec.

-------
            Open the control stopcock above the tube.   Open the  auxiliary
stopcock just long enough to clear bubbles from the tube.   Push  reset
button.  Bring the concentration index meter to 0.03.   (if  the concentra-
tion is above 0.03, add more electrolyte to dilute  to  0.03.   If  the  con-
centration is belov 0.03, add more subsamples from  Fraction 3 until  0.03
is reached.)  The final concentration should also be less than 10,000
particles/2 ml at manometer setting 15-2.
            Svitch back to time and open stopcock.   After pushing  reset,
wait k sec, then push accumulate.   When the calibration light comes  on,
push stop and close tube stopcock.
            Record the total count data from each channel.
            Drain residual electrolyte to a standard volume (130 ml) for
15-y tube analysis.  Transfer sample to a clean 250-ml storage beaker
rinsed with filtered electrolyte for analysis with  a 15-y tube.  Cover
storage beaker with cellophane to avoid contamination.   The 15-y tube
analysis should be completed within 2 hr of the 200-y  analysis.
            Change to the 15-y aperture tube and set the instrument
controls as follows:  active channel switch to lU-3, and gain control
to automatic.  Set the calibration potential and milliamp controls as
required.  Select the proper overlap channel with the  channel selector
switch.
            Pour the sample saved from the 200-y analysis through  a
clean 10-y micromesh sieve into a clean electrolyte beaker.   (The  sieve
should be washed with filtered electrolyte prior to use.)   Immediately
place the beaker on the sample stand and open the tube stopcock.  Open
auxiliary stopcock to clear bubbles from the tube and  push  the reset
button.
            Do not use the stirring motor.  Push accumulate button and
stop when the calibration light comes on.  Switch gain control to
manual and match difference percent from channel Ik with the difference
percent from channel 5 of the 200-y tube analysis.   Record  the data
from each channel.
                                3-35

-------
NOTE 1:  Always keep tinfoil shield door closed during analysis.
NOTE 2:  Make sure bubbles are clear from tube aperture by opening both
         stopcocks.
NOTE 3:  If tube clogs, brush aperture opening.  If tube is still clogged,
         clear vith an ultrasonic dismembrator.
NOTE h:  Always have aperture current in off position when not  running an
         analysis.
NOTE 5:  Check calibration and automatic gain control weekly.
Calculations
            Record the total weight of the sand (Fraction l) and the
silt plus clay (Fraction 2) fractions.  Weigh and record the weight of
each of the larger size fractions based on the dry sieving results
(Table 3-4).  The exact number of size fractions will depend on the
number of sieves used.  The percentage of each size fraction can be
calculated by dividing the weight on each sieve by the total weight
of the sample.
            Record the Coulter counter data in a form similar to
Table 3-5-  One fraction was counted with an aperture of 200 V.  These
channel counts are designated l6-A, 15-A, lU-A, etc.   (The number
refers to the channel number and the letter refers to the 200-y aperture
sample.)  Results for the 15-y aperture sample are designated l4-B,
13-B, etc.
NOTE:  When counting the 15-y aperture sample, the Coulter counter
       should be calibrated such that the 6-A reading is equal  to the
       lU-B reading.
            The numerical sum of three consecutive channels is  propor-
tional to one phi size.  Therefore, for the stated conditions,  the sum
of 16-A, 15-A, and lU-A is the U phi-size fraction.   The total  of 13-A,
12-A, and 11-A is the 5 phi-size fraction, etc.  These totals are desig-
nated size fraction A-H in Table 3-5 and are equivalent to phi  fractions
4-11.
            The following approach is used to calculate the percentage
of each phi fraction in the original sample:

                         % 4 phi =   .   . 100
                               3-36

-------
                               Table 3-1*
                Data Tabulation for Sand-Size Fractions
Sample No.
Analyst
Date
Sand Fraction (Fraction l)           	g
Sand and Clay Fraction (Fraction 2)  	g
                          Total Sample Weight


Coarse Fraction

j) Size   Dish #  Dish, vt, g  Dish and Sed, g  Cumulative vt, g  % Larger

  -3
  -2
  -1
   0
   1
   2

   3
   k
                               3-37

-------
                               Table 3-5
            Data Tabulation for Coulter Counter Results for
                    Silt- and Clay-Sized Fractions
Sample No.

Analyst

Date
           Sand Fraction

           Silt and Clay Fraction

           Total Sample Weight
  Size
Fraction
    A
    B
    D
    G
 Coulter
 Reading

  16-A
  15-A
  lU-A

  13-A
  12-A
  11-A
  10-A
   9-A
   8-A
   T-A
6-A/lU-B
  13-B
  12-B
  11-B
  10-B
   9-B
   8-B
   7-B
   6-B
   5-B
   U-B
   3-B
   2-B
   1-B
 Relative
Abundance
Phi
                                             10
                                             11
                               3-38

-------
where:
W = sum of three consecutive channels
X = total for size fractions A-H
Y = total weight of the silt and clay fraction in the original sample
    (Fraction 2), g
Z = total weight of the original sample (.silt and clay and sand), g
Method 2:  Sieving and Pipet Analysis
Apparatus
8-in. stainless steel 63-y sieve (wet sieve)
Distilled or demineralized water
Receiving container to fit under wet sieve, volume > 1000 ml
Drying oven
Mortar and pestle
8-in. stacking sieves:  7 at 1-phi intervals, -2 phi to +h phi; 1/2-phi
      intervals should also be available and can be used if desired.
Sieve shaker (Ro-Tap or equivalent)
Weighing dishes
Balance to 0.0001 g
1000-ml graduated cylinders
20-ml pipets with- controlled fill bulbs
Stirring rods
Constant temperature bath
50-ml preweighed beakers
Clock with second hand
Thermometer, in 1C divisions or better
Dessicator
Reagents
10 percent hydrogen peroxide, H20a-
1 percent Calgon:  dissolve 10 g commercially available Calgon in 1 Si
      of distilled water.
Procedure
            Homogenize the sample by mixing or mechanically tumbling.
Remove a UO- to 150-g subsample.  The smaller size is generally suitable
for fine-grained sediments while the larger size is needed when the

                                3-39

-------
particle sizes are veil distributed.  A flow diagram summarizing the
procedure is presented in Figure 3-^.   The next step is optional.  If
it is desired to determine the true sample particle-size distribution,
treat the sample with hydrogen peroxide as indicated to destroy organic
matter prior to sizing.
            If it is desired to determine the apparent particle-size
distribution, omit the treatment with hydrogen peroxide and proceed as
indicated.
            Place the sediment sample in a large beaker (>_ 2 &) and add
20 ml 10 percent hydrogen peroxide.  Let the sample stand until frothing
ceases and add an additional 10 ml hydrogen peroxide.  Continue the
incremental addition of hydrogen peroxide until no frothing occurs on
addition.
            Boil the sample to remove any excess hydrogen peroxide.
This should be completed in a large beaker to prevent sample loss due
to boiling over or frothing.
            Separate the sample into coarse and fine fractions by
wet sieving through a 63-y stainless steel sieve.  If possible, the
quantity of distilled or demineralized water used in the sieving
process should be kept below 900 ml.  Continue wet sieving until only
clear water passes through the sieve.   Collect the fine fraction that
passes through the sieve and retain the coarse fraction on the filter.
NOTE:  Never wet sieve using a brass sieve and always wet seive at
       room temperature.
            Coarse fraction.  Transfer the coarse fraction to a beaker
using tap water.  Dry the sample in an oven at a temperature not
exceeding 50C.  The temperature limitation is a precaution against
sample splattering or particles cementing together.
            Transfer dry sample to a dessicator for cooling.  Dis-
aggregate sample, if necessary, with a porcelain mortar and pestle.
            Transfer the dried sediment sample to a preweighed beaker
and determine the weight of the sediment sample.
            Build a nest of U. S. standard sieves of the required phi
sizes with, the coarsest sieve on the top and the finest sieve on the

-------
                                 SAMPLE
3A INCH ADD TO
COARSE FRACTION
                                   I
WET SIEVE 3M INCH
 MESH TO REMOVE
 LARGE FRACTION
COARSE FRACTION
      DRY
  DRY SIEVE
 EMPTY SIEVES
       i
WEIGH PORTIONS
     WET SIEVE
   No. 230 MESH
   PAN FRACTION
  FINE FRACTION
ADD PAN FRACTION
                               ADD PEPTISER
                                                           CHECK FOR
                                                         FLOCCULATION
                                                                llto
                                                        PIPETTE ANALYSIS
                                                               I
                                                        DRY WITHDRAWALS
                                                               I
                                                       WEIGH WITHDRAWALS
  Figure 3-^.     Particle-size procedure using sieving/pipet analysis
                                  3-Ul

-------
bottom.  Place a pan on the bottom, add the sample to the top sieve, and
place a lid on the nest of sieves.
            Place the nest of sieves on a Ro-Tap (or equivalent)  mechani-
cal shaker and shake for 10 to 15 min.  Empty each sieve onto a large
piece of paper.  Invert each sieve on a piece of paper, lightly tap the
screen, and brush the particles from the screen.  Do not touch screens
with your fingers.
NOTE:  Brushes should be of softer material than the screen material,
       i.e. steel brushes on sieve sizes of 1 phi or greater, brass or
       nylon, brushes on stainless steel screens, and nylon brushes on
       brass screens.
            Weigh, and record each size fraction.  If the weight on any
one sieve exceeds the value in Table 3-6, the sample should be recom-
bined, coned and quartered or split, and sieved again.
                               Table 3-6
                Maximum Sieve Loads on 8-in-diam. Sieves


Sieve Size
phi
-2.00
-1.00
0.00
+1.00
+2.00
+3.00
+U.OO
for 1-phi


mm
u
2
1
1/2
1A
1/8
1/16
Intervals

Maximum allowable retention
g
160
110
80
60
Uo
30
20
            The sum of all individual size fractions should be approxi-
mately equal to the original sample weight.  Sample losses and
inaccuracies should be less than 1 percent.
            Fine fraction.  Allow the fine fraction from the initial
sieving to stand until all silts and clays settle out.   Remove the
clear water by careful decantation or siphoning.   If, after 2^ hr,
sediment particles are still in suspension, measure the volume of
water decanted.  Sample the decanted water and determine the suspended
                             3-^2

-------
solids concentration.  Calculate the weight of sample lost (volume x
concentration) and correct results for this loss.
            Transfer the fine fraction to the metal cup of a malt  blender
and add 10 ml 1 percent Calgon solution.   The Calgon acts as a peptizer
to prevent the flocculation of sediment particles.   Mix the suspension
on a blender and transfer to a 1000-ml graduated cylinder.  Add distilled
or demineralized water to a volume of approximately 900 ml.   Mix.
            Let the sample stand for 2 to 3 hr and  observe for floccu-
lation.  If a definite clearing occurs, add 10 ml 1 percent Calgon and
repeat the process until no noticeable flocculation occurs.   Record the
volume of Calgon solution added.
            Dilute the sediment suspension to 1000  ml with either
distilled or demineralized water.  Thoroughly mix the sample.  Immedi-
ately withdraw a 20-ml sample from a depth of 20 cm and determine  the
wet weight of sediment withdrawn.  The total weight of sediment in the
graduated cylinder should be approximately 15 g and between 5 and  25 g.
If the total sample contains more than 25 g, a subsample should be used
for pipet analysis.  This can be obtained by pouring the complete
sample through a splitter trough or pouring off part of the well-mixed
sample and diluting the remainder.
            Place the graduated cylinder in a constant temperature bath
for the duration of the analysis as water viscosity, which varies  with
temperature, can affect sediment settling properties.  Immerse the
cylinder to the 1000-ml mark and firmly clamp in place for stability.
            Adjust the sample volume to 1000 ml, if necessary, and
thoroughly stir the sample.  Make sure that any settled sediment is
completely dispersed.
            About 15 sec after the stirring is stopped, insert a 20-ml
pipet to a depth of 20 cm.  (it is convenient to mark the stem of  the
pipet 10 and 20 cm from the tip.)  Withdraw the sample so the pipet is
full before 20 sec has elapsed since stirring was stopped.  Transfer
the sample to a preweighed 50-ml beaker.   Wash the  pipet with distilled
water and add the rinsing to the beaker.
            Withdraw 20-ml samples from a depth of  10 cm at the times

                             3-^3

-------
indicated in Table 3-7.  If necessary, samples may be collected at half
the indicated times and half the indicated depths.  Transfer the samples
to preweighed, 50-ml beakers.
            Dry the sample containing beakers at a temperature less than
100C.  Do not allow the samples to boil as this may result in loss of
samples.
            Transfer the beakers to a dessicator and cool.   Weigh samples
to the nearest 0.0001 g.
Calculations
            The data for both the coarse fraction and the fine fraction
should be recorded in tabulated form as shown in Table 3-8.  It should
be noted that the weights of the samples withdrawn during the pipet
analysis are automatically cumulative while those of the dry sieving
are not.  Also note that a correction for the peptizer must be included
in the pipet analysis results.  In addition, the calculation of the
percent finer and percent larger data uses the total sample weight
(i.e. weight of fine fraction plus the coarse fraction).
            The total weight of the fine fraction is determined from
the sampling of the peptized sediment suspension.   The total weight of
each fine fraction is calculated by multiplying the sample  weight by
50.  If the initial sample volume is something other than 1000 ml and/or
the sample size is something other than 20 ml, this factor  must be
appropriately corrected.

-------
                                              Table 3-7
                             Sampling Time Intervals for Pipet Analysis
                               Elapsed time for withdrawal of sample in hours (h.), minutes (m), and
                               	seconds (s)    	
Diameter Diameter
  finer    finer  Withdrawal
  than     than      depth
   phi	M	cm      18C   19C   20C   21    22C   23C   2UC   25C   26C   27C
ll.O
I..5
5.0
5.5
6.0
7.0
8.0
9-0
10.0
11.0
62.5
UU.2
31.2
22.1
15.6
7.8
3.9
1.95
0.98
O.U9
20
20
10
10
MO
10
5
5
5
5
20s 20s 20s 20s 20s 20s 20s 20s 20s 20s
2mOs Im57s lm5Us Im51 lmU9s lnA6s ImUiis ImUls Im39s Im37s
Restir Restir Restir Restir Restir Restir Restir Restir Restir Restir
2mOs Im57s Im5^s Im51s lmU9s lmU6s Im^s Irakis Im39s Im37s
UmOs 3m5Us 3mlt8s 3mU2s 3m37s 3m32s 3m27s 3m22s 3ml8s 3ml3s
8mOs 7mU8s 7m36s 7m25s 7ml5s 7m5s 6m55s 6m*i5s 6m36s 6m27s
31m59s Slmlls 30m26s 29mUls 28m59s 28ml8s 27m39s 28mls 26m25s 25mU9s
63m58s 62m22s 60m51s 59m23s 57m58s 56m36s 55ml8s 5^m2s 52mU9s 51m39s
Uhl6m Uh9m Uh3m 3h58m 3h52m 3hU6m Shiilm 3h36m 3h31m 3h27m
17h3m l6h38m l6hlUm 15h50m 15h28m 15h6m lUhU5m lUh25m lUh5m 13hi+0m
68hlUm 66h32m 6Uh5^m 63h20m 60h50m 60h23m 58h59m 57h38m 56h20m 55h5m

-------
                               Table 3-8
             Typical Particle-Size Distribution Data Sheet
Sample No.
Analyst
Date
Total Sample Weight = Coarse
Fine Fraction (Pipette Analysis)
Peptizer (l% Calgon) added 	ml
Peptizer Correction Factor
Water Bath Temperature 	
Fine Fraction
                           g + Fine
                     wt/20 ml Sample =
                     C
                Dish vt  Dish and Sed  Less Peptizer  Total wt
4) Size  Dish #     g     	g       	g	      g     % Finer
   U
   5
   6
   7
   8
   9
Coarse Fraction
                  Initial vt
                                           _g
  Size
   -5
   -U
   -3
   -2
   -1
    0
    1
    2
    3
    U
Dish  Dish wt  Dish and Sed  Cumulative wt
  #      g     	g	  	g	  % Larger  Comments
                                 3-U6

-------
                              References
1.  U. S. Geological Survey.   "National Handbook of Recommended Methods
    for Water Data Acquisition."  Office of Water Data Coordination,
    Geological Survey, U.  S.  Department of the Interior;  Reston,
    Virginia (.1977).
2.  Guy, H. P.  "Techniques of Water-Resources Investigation  of the
    United States Geological Survey."  Chapter Cl.   "Laboratory Theory
    and Methods for Sediment Analysis."  59 p. (1973).
3.  Fochtman, E. G.  "Selection of a Particle Size Measurement
    Instrument."  In:  Particle Size Analysis; Stockman,  J. D., and
    Fochtman, E. G. (.Eds.),  pp. 125-129. .Ann Arbor Science  Publ.
    ikO p. (1978).
h.  Walton, A.  "Methods for Sampling and Analysis of Marine  Sediments
    and Dredged Material."  Scientific Information and Publications
    Branch, Fisheries and Marine Service, Department of Fisheries
    and the Environment; Ottawa, Ontario, Canada.   7^ p. (1978).

-------
                                  PH
            The pH of a solution is a measure of the hydrogen ion
activity.  It is mathematically expressed as the negative logarithm of
the hydrogen ion concentration.  This parameter is important because it
is used to calculate the species distribution of weak acids and bases
such as carbonates, ammonia, and cyanide that may be present in the
sample.  Acid-base equilibria is important because the toxicity of a
sample can vary with the distribution of chemical species.   The recom-
mended analytical method is electronic measurement using glass elec-
trodes.

                      Sample Handling and Storage

            Whenever possible, pH measurements should be taken and
recorded in situ.  If necessary, the sample can be returned to the
laboratory in either glass or plastic containers.  However, the
measurements should be completed as soon as possible as there is no
known method of preserving sample pH.  It is also recommended that
sample containers be completely filled and tightly sealed when pH
analyses are to be run on a delayed basis.  This precaution is
intended to minimize the exchange of carbon dioxide that can alter
pH.  Additionally, sample should be kept in the dark and refrigerated.

                          Sample Preparation

            The only required preparation is to ensure that the
samples to be measured and the standard pH buffers are equilibrated
at the same temperature.
                                3-H8

-------
              Procedures for Water Samples (Wl, W2? S1A)

Method 1:  Glass Electrode
Apparatus
Electronic pH meter:  a meter with a temperature compensation adjust-
            ment vould be preferred but not required
Glass electrode:  if pH values greater than 10 and high sodium
            concentrations are anticipated, minimum sodium error
            electrodes should be selected
Reference electrode such as the saturated calomel capable of main-
            taining a constant potential
Magnetic stirrer and stirring bars
Reagents
Standard pH buffers throughout the anticipated range of sample pH
            values.  Prepared buffer solutions and dry powders are
            available commercially.  For more accurate work, fresh
            buffer solutions can be prepared as needed.
Procedure
            Follow manufacturer's directions to set up and warm up the
pH meter.  Adjust the temperature compensator to the temperature of
the samples and buffers.
            Select two buffer solutions in the approximate range of
the samples to be measured.  Place the first buffer in a beaker and
stir gently on a magnetic stirrer.  Lower the electrodes in the buffer
and allow the meter needle to stabilize.  Adjust the instrument cali-
bration control to the correct buffer value.
            Remove the electrodes from the buffer solution and rinse
with distilled water.   Gently dry the electrodes with soft, absorbent
tissue.
            Place the second buffer solution in a beaker and stir
gently on a magnetic stirrer.  Lower the electrodes in the buffer and
allow the meter needle to stabilize.  The reading should be within
0.1 pH units of the expected buffer value.
            Remove the electrodes from the buffer, rinse, and dry
as before.
            Transfer 50 to 100 ml of a sample to be measured to a
beaker and mix with a magnetic stirrer to ensure homogeneity.   Immerse

                                3-1*9

-------
the electrodes in the sample, allow the instrument to stabilize, and
record the pH.  It is a good practice to simultaneously measure the pH
and the temperature of the sample "because temperature affects dissocia-
tion constants of acids and bases and, hence, the significance of pH.
            Continue processing samples, making sure to rinse and dry
the electrodes between samples.  Buffer solutions should be run approxi-
mately once an hour to check the instrument standardization.
                              3-50

-------
                 Procedures for Sediment Samples (SIX))

Method 1:  Glass Electrode
Apparatus
Electronic pH meter:  a meter with a temperature compensation adjust-
            ment vould be preferred but not required
Glass electrode:  if pH values greater than 10 and high sodium con-
            centrations are anticipated, minimum sodium error
            electrodes should be selected
Reference electrode such as the saturated calomel capable of main-
            taining a constant potential
Magnetic stirrer and stirring bars
Reagents
Standard buffers throughout the anticipated range of sample pH values.
            Prepared buffer solutions and dry powders are available
            commercially.  For more accurate work,  fresh buffer
            solutions can be prepared as needed.
Procedure
            Standardize the pH meter as described for water samples.
            Transfer an aliquot of blended, moist sediment sample to
an appropriate sized beaker.  Do not use dried or frozen sediment
samples as the dehydration process is not known to be reversible.
Insert the electrodes in the sample and allow the instrument to
equilibrate.  Record the pH and the temperature of the sample.
            Clean the electrodes and process the next sample.   Check
the standardization of the pH meter at regular intervals using known
buffer solutions.
            If the sample is sufficiently dry that  a direct pH
reading cannot be taken, slurry the sediment with a known volume of
distilled water.  Report the pH of the slurry and the solid-liquid
ratio of the slurry.
                                 3-51

-------
                     OXIDATION REDUCTION POTENTIAL

            The oxidation-reduction potential (redox potential or Eh)
is defined as the electromotive force developed by a platinum electrode
immersed in a vater or sediment sample relative to a standard hydrogen
electrode or a reference electrode of known Eh.  The obtained value is
a crude estimate of the oxygen status of the sample.  A positive value
indicates that the water or sediment sample is in an oxidized state or
oxygen is present.  A negative value would indicate an absence of oxygen
or reducing conditions.

                     Sample Collection and Storage

            The preferred method of obtaining oxidation-reduction
potential data is in situ measurement.  If this is impractical, the
measurements should be made as soon as possible.  Since exposure to
the atmosphere may affect the oxidation-reduction potential of the
sample (oxygen may dissolve in water or oxidize sediments), pre-
cautions should be taken to minimize sample contact with the atmosphere
prior to measurement of the oxidation-reduction potential.   This
precaution will necessitate the use of wet sediment samples for the
measurement.

      Procedure for Water (Wl, W2, S1A) and Sediment Samples (SID)

Method 1:   Platinum Electrode
Apparatus
Potentiometer or pH meter equipped to read in millivolts
Saturated potassium chloride calomel cell
Platinum electrode with clean platinum surface
Procedure
            Insert the platinum electrode into the Wl water sample or
the SID sediment sample and allow the instrument to equilibrate.
Record the instrument millivolt reading.
                                3-52

-------
Calculations
            Sample potential = (measured millivolt  reading)
                    - (standard calomel  potential)
                               3-53

-------
                   TOTAL SOLIDS AND VOLATILE SOLIDS

            Total solids or total residue refers to the material
remaining after a sample has been dried or evaporated at a specific
temperature.  The residue, therefore, includes both soluble and
suspended solids in the original sample.  The procedure should be
considered operationally defined to the extent that slightly different
results can be obtained if samples are dried at temperatures other
than those specified.  These differences would be expected to be more
pronounced for sediment samples than for water samples.
            Volatile solids procedures have been included in this
section because the residue from the total solids determination is
used as the starting material.  The solids are subjected to ignition
at a higher temperature to provide a crude estimate of the organic
matter in the total solids.  The use of temperature to distinguish
between organic and inorganic solids, tiowever, should be used with
caution because some organic material can be lost at the lower drying
temperature and some inorganic material (carbonates and chlorides) can
be lost at the higher ignition temperatures, particularly with sediment
samples.

                      Sample Handling and Storage

            The suggested method of handling total solids and total
volatile solids samples is presented in Figure 3-$.  Only moist
sediment samples should be used for this determination.  Either glass
or plastic containers can be used for storing the samples.  However,
the storage period should be kept to a week or less.

-------
vn
VJ1
                         WATER SAMPLE
                                                                   DREDGE SAMPLE
1

r
STORE
I
DESSICATE
1

+
FILTER
I
STORE
I
DESSICATE



^^
NO TREATMENT
(W3)

__
                                                                    STORE WET
         PURPOSE
         CONTAINER
         SAMPLE TREATMENT
         PRESERVATIVE
         STORAGE TIME
           Figure  3-5.
                                   Total Solids  Soluble or
                                      or        Flltrable
                                  Total Residue    Residue
                                      G,P
                                     None
                                                  G,P
                                                 Filter
                   /."C
                            Used in
                           Elutriate
                              G,P
                                                            None
Mobile
 Cone.
 G,P
                                                                        None
                                                                                                              CORE SECTION
SAMPLE
DESIGNATION
Wl
W2
W3
SIA
SID
Percent
 Solids
 G.P
                                                                                                           None
                                                                           (Minimize Air Contact.  Keep Field Moist )
        Iw          Iw         Iw          Iw

Handling and storage  of  samples for total  solids  and volatile solids analysis

-------
                               Total Solids
                 Procedure for Water Samples (Wl,  W2, S1A)

Method 1;   Gravimetric
Apparatus
Evaporating dishes of 100-ml capacity, either porcelain,  platinum,  or
            Vycor
Muffle furnace for operation at 550 +_ 50C
Steam bath or drying oven
Desiccator
Analytical "balance
Procedure
            Ignite a clean evaporating dish at 550 +_ 50C for 1 hr  in
a muffle furnace.  Cool, desiccate, and weigh the  dish.   Record the
weight of the empty dish and store the evaporating dish  in a desiccator
until used.
            Transfer a known volume of Wl, W2, or  S1A sample to the
preweighed dish and evaporate to dryness on a steam bath or in a drying
oven.  It is recommended that a sample size be chosen that will produce
a minimum residue of 25 mg.  With low residue waters, successive
aliquots of the sample should be added to the same evaporating dish
until the required minimum residue is obtained.
            Exclude large, nonhomogeneous materials from the sample.
Also, uniformly disperse any floating oil and grease before subsampling.
            When drying in an oven, evaporate the  sample at 98C.   This
precaution is necessary to prevent loss of sample  by boiling and
splattering and, hence, low results.
            After evaporation, increase the drying oven  temperature
from 98C to 103 to 105C or transfer the evaporating dish from the
steam bath to a drying oven set at 103 to 105C.   Dry the sample for
1 hr at 103 to 105C.  Cool, desiccate, and weigh the sample.  Repeat
the 1-hr drying cycle at 103 to 105C until a constant  weight is
obtained for the residue.  The gain in weight of the tared evaporating
dish is a measure of the solids or residue of the  sample.
                                 3-56

-------
Calculations
            The solids concentration of the sample is calculated, "by
dividing the weight of the residue by the volume of sample used:
                        ,     . ,     (A- B) x  loop
                      mg/8 residue =	
where
     A = weight of dish and sample residue* mg
     B = weight of dish, mg
     V = volume of sample aliquot, mC
            When a 1 sample is used,  the results should be termed
total solids or total residue. When a W2 or SI sample is used,  the
results should be termed total filterable solids or total filterable
residue.
                                3-57

-------
                              Total  Solids
                   Procedure  for  Sediment  Samples  (.SID)

Method 1:  Gravimetric
Apparatus
Evaporating dishes of 100-ml capacity, either porcelain, platinum, or
            Vycor
Muffle furnace for operation at 550 +_ 50C
Steam bath or drying oven
Desiccator
Analytical balance
Procedure
            Ignite clean evaporating dishes at 550 +_ 50C for 1 hr in
a muffle furnace.  Cool, desiccate, and veigh each dish.  Record the
weight of each dish and store the dishes in a desiccator until used.
            Homogenize an SID sample and transfer a 25-g aliquot to
a tared evaporating dish.  Weigh the sample-containing dish to the
nearest 10 mg.  Dry the sample overnight in a drying oven at 103 to
105C.  Cool the sample, desiccate, and weigh the sample.  Repeat the
drying process until a constant weight residue is obtained.
Calculations
            The total solids or total residue of the sediment samples
are calculated by dividing the weight of the dried residue by the initial
weight of the sample.  Results are termed % solids:
                        % Solids = 4 x 10
                                   (_> * _D
where
     A = weight of dish and dry sample residue
     B = weight of dish
     C = weight of dish and wet sample
                                 3-58

-------
                     Volatile Solids Determination

            Volatile solids are determined by ashine the dried residue
from the total solids, filterable solids, or percent solids determination
at 550 +_ 50C.  The weight of material lost at the higher temperature is
normalized to the initial volume or weight of sample and reported as
percent volatile solids.
Apparatus
Evaporating dishes of 100-m8 capacity, either porcelain, platinum, or
            Vycor
Muffle furnace for operation at 550 + 50C
Desiccator
Analytical balance
Procedure
            Preheat a muffle furnace to 550 +_ 50C.  Ignite the residue
from the total solids (Wl), filtrable solids (W2, S1A), and/or percent
solids (SID) determinations to a constant weight.
            Remove the samples from the furnace and allow them to
partially cool.  Transfer the samples to a desiccator for final cooling.
Weigh the sample dishes as soon as they are cool.
Calculations
            The material lost on ignition is referred to as volatile
solids or volatile residue.  The material retained in the evaporating
dish is referred to as fixed solids or fixed residue.
Wat er Sample s:
                  it,  '-i 4.-n     *     (A - D) x lOOQ
                mg/x. volatile residue = 		
                    /n -P-  A    -A     (D - B) x loop
                  mg/ fixed residue =	
where
     A = weight of dish and dry sample residue
     D = weight of dish and ignition residue
     V = volume of original sample, m2
     B = weight of evaporation dish
                                3-59

-------
Sediment Samples:
                                        A - D
                     volatile residue = - -   x 100
                                       T\   -D
                     % fixed residue =   ~   x 100
where
     A = weight of dish and dry sample residue
     B = weight of evaporation dish
     D = weight of dish and ignition residue
                                3-60

-------
                           SPECIFIC GRAVITY

            The specific gravity of a substance is defined as the ratio
of the mass of a given volume of the substance to an equal volume of
distilled water at the same temperature.  Since the specific gravity of
water is 1 g/cc, the specific gravity of the solid is equivalent to the
grams of dry solid/cc.

                      Sample Handling and Storage

            Since specific gravity is equivalent to the mass of dry
solids per unit volume, sediment samples for this determination may
be stored either wet, dried, or frozen.

             Procedure for Sediment Samples (S1A, S2, S3)1

Apparatus
Constant volume pycnometer
Drying oven
Thermometer
Balance
Distilled water
Procedure
            Fill pycnometer with distilled water and weigh to the
nearest 0.1 mg.  Record the temperature  of the water.
            Weigh a sample of oven-dried sediment.  Remove a small
amount of water from the pycnometer and  add the known mass of dried
sediment to the pycnometer.
            Apply a suction or boil the  suspension to remove any air
bubbles that may have been introduced into the pycnometer with the
sediment sample.  If the sample is boiled, cool the sample to the same
temperature recorded for distilled water.
            Weigh the pycnometer filled  with the sediment suspension
and record the weight to the nearest 0.1 mg.
                                3-61

-------
Calculations


            Calculate the specific gravity of the sediment sample as


follows:

                                    W

                          G  =
                           s     -    + 
                                w    ws    s
where
      G  = specific gravity of the sample
       S



      W  = mass of sediment used, g
       b>



      W  = mass of the water-filled pycnometer, g




        = mass of the pycnometer filled with a water-sediment

           suspension, g
                                 3-62

-------
                              References
1.  U. S. Geological Survey.   "National Handbook of Recommended Methods
    for Water-Data Acquisition."  U.  S. Geological Survey,  U.  S. Depart-
    ment of the Interior; Reston,  Virginia (1977).
                                3-63

-------
INORGANIC ANALYSIS

     Carbon
          Organic
          Inorganic
     Metals
          Aluminum
          Cadmium
          Calcium
          Chromium
          Copper
          Iron
          Lead
          Magnesium
          Manganese
          Molybdenum
          Nickel
          Zinc
          Arsenic
          Mercury
          Selenium
     Nitrogen
          Ammonia
          Nitrate
          Nitrite
          Total Kjeldahl
          Organic
     Phosphates
          Soluble Reactive
          Total
          Organic
     Sulfides

-------
                  CARBON, TOTAL ORGANIC AND INORGANIC

            Carbon may exist in sediment and water samples as either
inorganic or organic compounds.  Inorganic carbon is present as carbo-
nates, bicarbonates, and possibly free carbon dioxide.  Specific types
of compounds that are considered to be included in the organic carbon
fraction are nonvolatile organic compounds (sugars), volatile organic
compounds (mercaptans), partially volatile compounds (oils), and
                                                  #
particulate carbonaceous materials (cellulose).1*2
            The basis of the method is the catalytic or chemical
oxidation of carbon in carbon-containing compounds to carbon dioxide
followed by the quantification of the carbon dioxide produced.
Alternately, the carbon may be reduced to methane and appropriately
quantified.   It follows, then, that the distinction between inorganic
carbon and organic carbon is the method of sample pretreatment.   There
are presently two procedures for defining this separation.  One method
is based on sample treatment with a strong acid.  Analysis of an
untreated sample is a measure of total carbon while analysis of the
acid-treated fraction is a measure of organic carbon.  Inorganic carbon
is calculated by subtraction.  The second method of separation is
based on differential thermal combustion with organic compounds being
converted to carbon dioxide at 500C to 650C3''* and inorganic carbon
being converted to carbon dioxide at 950C to 1300C.'*'5

                      Sample Handling and Storage

            Flowcharts for the handling of samples intended for organic
carbon and inorganic carbon analysis are presented in Figure 3-6 and
Figure 3-7.   Water and sediment samples to be analyzed for inorganic
carbon may be stored in glass or plastic containers.  There is no
effective preservative because of the carbon dioxide reserve in the
atmosphere.   The only precaution that can be taken for inorganic

* References for this procedure can be found on page 3-76.

                                 3-65

-------
o\
cr\
                                                                                                   CORE SAMPLE
AC 1 D 1 FY
1
r
STORE

[DREDGE SAMPLE
|
^
r

CORE SECTION
1
NO TRE
(




&TMENT
rf3)












ELUTRIATE






STOR
i



EWET |
r
^
DIGEST

                I
                                                                                                       1
SAMPLE DESIGNATION
PURPOSE
Wl
Total Water
Cone.
W2
Soluble
Water
Cone.
W3
Used In
Elutriate
SIA SIB
Mobile
Cone.
SIC S1D S2
Total
Sediment
Cone.
S3

              CONTAINER
              SAMPLE TREATMENT
              PRESERVATIVE
             .STORAGE TIME
              DIGESTION SOLUTION
             .SAMPLE VOLUME  OR WEIGHT
                                         G,P
                                         None
                                                    G,P
                                                    Filter
                    G,P
                                                                None
                                                                            G,P
                                                                            None
                                                                                                               :5.P
                                                                                                               None
                                          hr
(Minimize Air Contact.)





        2*1 hr
                                                                
-------

WATER SAMPLE ORE
f
* t *
ACIDIFY FILTER N TIT JT
1 1 	
STORE ACIDIFY

*
STORE 1 | ELUTRIATE
1 I
ANALYZE ANALYZE ANALYZE
(Wl) (W2) (S1A)

S SAMPLE DESIGNATION Wl W2 W3
-g
PURPOSE Total Water Soluble Used in
Cone. Water Elutriate
Cone.
CORE SAMPLE
j


*
DGE SAMPLE CORE SECTION
1



ORE WET | DRY
, 1
STORE
, ,


ANALYZE ANALYZE
(SID) (S2)

S1A SID
Mobile Total
Cone. Sediment
Cone.
S2
Total
Sediment
Cone.


FREEZE
I
STORE

ANALYZE
(S3)

S3
Total
Sediment
Cone.
CONTAINER
SAMPLE TREATMENT
PRESERVATIVE
STORAGE TIME
SAMPLE VOLUME OR WEIGHT
                           None
                                       Filter      None
                         HC1  or H2SO,,  HC1 or H2SO,,    1|C
                           to pH2       to pH2
                                                               G,P
                                                               None
<4C
                              hr
                                         hr
                         30-100 ml     30-100 ml     Variable
                                   G,P
G,P
                                                                                                  None     Air dry
                                             None
G,P
                                                                                                                      Freeze
                                                        None
                                0.5-5.0 9  0.5-5.0 g   0.5-5.0 g
          Figure  3-7.      Handling  and storage of samples  for  total  organic carbon analysis

-------
carbon is to completely fill the sample container at the time of
sampling (exclude all air "bubbles), tightly seal the container,  and
complete the analysis immediately (Figure 3-6).
            Water samples for organic carbon analysis should be  stored
in glass containers unless substitute containers have been shown not
to affect total organic carbon (TOG) analyses.   Samples should be
processed as soon as possible (within 2h hr if  possible) to minimize
change due to chemical or biological oxidation.   Atmospheric uptake
of carbon dioxide is less critical since it would be evolved when the
sample is acidified prior to analysis.  Sediment samples for organic
carbon analysis may be stored in either plastic  or glass containers
(Figure 3-7).  Air drying of sediments (S2) may lead to low TOC
results due to oxidation or volatilization.  Therefore, moist storage
(SID) or frozen storage (S3) would be the preferred method of storage.
If samples are frozen, excessive temperatures should not be used to
thaw the samples.
                                 3-68

-------
               Procedure for Water Samples (Wl, W2, S1A)


Method 1:  Infrared Analysis6'7

Apparatus

Sample homogenizer such as a Waring blender or ultrasonic blender

Magnetic stirrer

Hypodermic syringe

Total carbon analyzer, either a single channel or a dual channel
            instrument  (Dow-Beckman Carbonaceous Analyzer Model
            No. 915, Dohrmann Envirotech DC-50 carbon analyzer,
            Oceanography International Total Carbon Analyzer, Leco,
            or equivalent)
Reagents

Distilled water:   the distilled water used in the preparation of
            standards and dilution of samples should be of the
            highest quality in order to have  a small blank.

Organic carbon, stock solution, 1000 mg/ C:   dissolve 2.125 g
            anhydrous potassium biphthalate,  KHCs Hi0u,  in distilled
            water and dilute to 1 & in a volumetric flask.

Organic carbon, standard solutions:   prepare  standard solution by
            dilution of the stock solution as required.

Inorganic carbon, stock solution, 1000 mg/C:   dissolve 3.500 g sodium
            bicarbonate, NaHCOs,  and i;.Hl8 g  sodium carbonate, NaaCOs,
            in distilled water in a 1-& volumetric flask and make up
            to the mark.

Inorganic carbon, standard solution:   prepare standards from the stock
            solution as required.

Packing for total carbon tube:   dissolve 20 g cobalt nitrate,
            CO(N03)2 ' 6H20, in 50 ml distilled water.  Add this
            solution to 15 g long-fiber asbestos in a porcelain
            evaporating dish.  Mix and evaporate to dryness on a
            steam bath.  Place the dish in a  muffle furnace and
            bring to 950C.  After 1 to 2 hr  at this temperature,
            remove the dish and allow to cool.   Break up any large
            lumps and mix adequately but not  excessively.  With the
            combustion tube held in a vertical position, taper joint
            up, put about 1/2 in.  of untreated asbestos in the tube
            first, then transfer in small amounts, approximately 1 g
            of catalyst into the tube with forceps or tweezers.  As
            it is added, tap or push the material gently with a lA-in.
            glass rod.  Do not force the packing.  The weight of the
            rod itself is sufficient to compress the material.  When
            completed, the length of the packing should be about 5 or
            6 cm.  Test the packed tube by measuring the flow rate of


                                 3-69

-------
            gas through it at room temperature,  and then at  750C.
            The rate should not drop more than 20 percent.
Packing for carbonate tube (dual channel instrument):   place a small  wad
            of quartz wool or asbestos near the  exit end of  the
            carbonate evolution tube.  From the  entrance end add 6  to
            12 mesh quartz chips, allowing these to collect  against the
            wad to a length of 10 cm.  Pour an excess of 85  percent
            phosphoric acid, HaPOi*, into the tube while holding it
            vertically and allow the excess to drain out.
Nitrogen gas, carbon dioxide free.
Procedure
            Turn on the infrared analyzer, recorder, and tube furnaces,
setting the total carbon furnace at 950C and the carbonate  furnace at
1T5C.  Allow sufficient warm-up time for stable, drift-free operation;
about 2 hr is required.  If used daily, the analyzer can be  left on
continuously.  Adjust the oxygen flow rate to 80 to 100 ml/min through
the total carbon tube.  With other instruments,  follow manufacturer's
directions to warm up the instrument*.
            Immediately prior to carrying out calibrations or analyses,
inject several portions of the appropriate standard into the tube to  be
used, until constant readings are obtained.   The actual injection
technique is as follows:  rinse the syringe several times with the
solution to be analyzed, fill, and adjust the volume to be pipeted.
Wipe off the excess with soft paper tissue, taking care that no lint
adheres to the needle.  Remove the plug from the syringe holder, insert
the sample syringe, and inject the sample into the combustion tube  with
a single, rapid movement of the thumb.   Leave the syringe in the holder
until the flow rate returns to normal,  then replace it with  the plug.
            Successively introduce a convenient  sized aliquot (20 to
50 yl) of each organic carbon standard and a blank into the  total carbon
tube and record peak heights.  Between injections allow the  recorder
pen to return to its baseline.  When a dual channel instrument is used,
the standardization procedure must be repeated using carbonate standards
to calibrate the low temperature channel.
            Thoroughly mix the sample.   Inject a convenient  sized
aliquot (20 to 50 yl) of the sample into the total carbon tube and
                                3-70

-------
record the peak height.  This result is a measure of the organic  carbon
concentration and the inorganic carbon concentration of the sample.
            Thoroughly mix the sample using a Waring blender or an ultra-
sonic homogenizer.  Transfer 10 to 15 ml of sample to a 30-ml beaker and
acidify with concentrated HC1 to a pH of 2 or less.  Purge the sample
with carbon dioxide free nitrogen gas for 5 to 10 min.   Plastic tubing
should not be used during the purging process unless it has been
previously shown that it will not add organic carbon to the sample.
            Mix the acidified sample on a magnetic stirrer.  While
stirring, withdraw a subsample from the beaker using a hypodermic
needle with a 150-um opening.  Inject the sample into the carbon
analyzer to be used and record the peak height.  This result is a
measure of the organic carbon concentration of the sample.
            Using either clear or filtered water samples, analytical
precision will approach 1 to 2 percent or 1 to 2 mg/1 carbon, whichever
is greater.  Analytical precision for unfiltered water samples will
increase to 5 to 10 percent because of the difficulty associated with
sampling particulate matter and the fact that the needle opening of
the syringe limits the maximum size of the particles that can be
included in the sample.
Calculations
            Dual-channel instrument.  Prepare calibration curves
derived from the peak heights obtained with the standard total carbon
and inorganic carbon solutions.
            Determine the concentration of total carbon and inorganic
carbon in the sample by comparing sample peak heights with the cali-
bration curves.
            Determine the concentration of total inorganic carbon in
the sample by subtracting the organic carbon value from the total
carbon value.
           , Single-channel instrument.  Prepare a calibration curve
derived from the peak heights obtained with the standard total carbon
solutions.  Determine the total carbon concentration in the sample by
comparing the peak height of the first sample injection with the
                                3-71

-------
calibration curve.   Determine the organic  carbon  concentration  in the
sample by comparing the peak height  of  the second sample  injection with
the calibration curve.   Inorganic carbon concentrations are calculated
by subtracting the organic  carbon concentration from the  total  carbon
c one entrat i on.
                                3-72

-------
              Procedures for Sediment Samples (SID,  S3)

Method 1:  Sample Ignition
Apparatus
Induction furnace such as the Leco WR-12,  Dohrmann DC-50,  Coleman CH
            analyzer, or Perkin Elmer 2UO elemental  analyzer
Combustion boats
Microbalance
Desiccator
Reagents
10 percent hydrochloric acid:  mix 100 ml concentrated HC1 with 900 ml
            distilled water.
Copper oxide fines.
Benzoic acid.
Procedure
            Dry at TOC and grind the sediment sample.
            Weigh a combustion boat and record the weight.  Place 0.2
to 0.5 g homogenized sediment in the combustion boat and reweigh.
Combustion boats should not be handled with the bare hand  during this
process.
            If total carbon or inorganic carbon is to be determined,
Cupric oxide fines may be added to the sample to assist  in combustion.
Combust the sample in an induction furnace.  Record  the  result as total
carbon.
            If organic carbon is to be determined, treat a known weight
of dried sediment with several drops of 10 percent HC1.  Wait until
the effervescing is completed and add more acid.  Continue this process
until the incremental addition of acid causes no further effervescence.
Do not add too much acid at one time as this may cause loss of sample
due to frothing.
            Dry the sample at TOC and place in a desiccator.  Add
Cupric oxide fines, combust the sample in an induction furnace, and
record the result as organic carbon.
                                3-73

-------
Calculations
            The carbon content of the sample can be calculated as:
              ,*


-------
      g = weight of sample combusted, g
            The organic carbon, CQ, concentration of the sample (in mg/g)
is calculated as follows:
                            C    (Xo)
                              "    (g)
where
     x
      o = weight of COa evolved at 650C,  mg
      g = weight of sample combusted, g
            Inorganic carbon, C   (in mg/g)  is calculated as:
                             CT   C.  - C
                              I =  t    o
Method 3:  Wet Combustion4'8
            A third method has been used for carbon in sediments.   This
is based on the oxidation of the sample with dichromate and back titra-
tion of the sample with ferrous ammonium sulfate.   References are
provided for the procedure but details are not given.   The procedure
is similar to the chemical oxygen demand test which is not specific
for carbon.  The wet combustion method is a redox  procedure and any
reduced chemicals in the sediment samples (ferrous iron, manganous
manganese, sulfide) will react with the dichromate.  Therefore, this
procedure is not recommended unless other instrumentation is not
available.
                                3-75

-------
                              References

1.  U. S. Environmental Protection Agency.   "Manual of Methods for
    Chemical Analysis of Water and Wastes."  Methods Development and
    Quality Assurance Research Laboratory,  National Environmental
    Research Center; Cincinnati, Ohio.  298 p. (197*0.

2.  U. S. Environmental Protection Agency.   "Methods for Chemical
    Analysis of Water and Wastes."  Environmental Monitoring and
    Support Laboratory, Office of Research  and Development, SPA;
    Cincinnati, Ohio (1979).

3.  Giovannini, G., Poggio, G., and Sequi,  P.  "Use of an Automatic
    CHN Analyzer to Determine Organic and Inorganic Carbon in Soils."
    Unpublished Report, Laboratory of Soil  Chemistry, via Corridoni,
    Pisa, Italy.  9 p. (1975).
k.  Konrad, J. G., Chesters, G., and Keeney, D. R.  "Determination of
    Organic- and Carbonate-Carbon in Freshwater Lake Sediments by a
    Microcombustion Procedure."  J. Thermal Analysis 2:199-208 (1970).

5.  Kemp, A. L. W.  "Organic Matter in the  Sediments of Lakes Ontario
    and Erie."  Proc. 12th Conference Great Lakes Research 12:237-2^9
    (1969).
6.  Environment Canada.  "Analytical Methods Manual," Inland Waters
    Directorate, Water Quality Branch; Ottawa, Canada (197*0-
7-  American Public Health Association.   Standard Methods for the
    Examination of Water and Wastewater. APHA; New York, New York.
    1193 p. (1976).
8.  Gaudette, H. E., Flight, W. R., Toner,  L., and Folger, D. W.
    "An Inexpensive Titration Method for the Determination of Organic
    Carbon in Recent Sediments."  J. Sed. Petrology Mt:2**9-253 (197*0.
                                3-76

-------
                                 METALS
           (Al, Cd, Ca, Cr, Cu, Fe, Fb, Mg, Mn, Mo, Ni, Zn)

             Metals are naturally occurring elements that distribute
themselves among several different chemical forms.  These forms include
dissolved metals, soluble metals, complexed metals, and particulate metals.
The actual distribution between these forms will depend upon factors such
as pH, redox potential, the presence of complexing molecules, and the
specific environmental chemistry of each metal.1
             Metals may reach waterways and, hence, sediments, as a result
of erosion and/or weathering of geological formations.  In addition,
significant quantities of metals are mobilized by man as the result of
mining, milling, lumbering, and similar activities.2  Once in a waterway,
metals are of concern because they may be essential in low concentrations
to the growth of some organisms;3 they may bioaccumulate to undesirable
levels in some organisms;  or they may be acutely toxic at high concentra-
tions. 3  The actual impact of metals will be a function of the metal
concentration, the distribution of the metal between the various chemical
forms, and the sensitivity of the organisms exposed.
             The most universally available instrument for the analysis
of a wide spectrum of metals is an atomic absorption spectrophotometer.5'6
This instrument provides a method that is relatively free from spectral
or radiation interferences because each metal has its own characteristic
absorption wavelength.  In addition, the method is more sensitive than
flame photometry or most colorimetric determinations.  The sensitivity
of atomic absorption spectrophotometry can be further extended by graphite
furnace atomization or sample concentration by chelation-extraction.
Therefore, atomic absorption is the recommended method of metal analysis.
Because of the similarity in the method of analysis, aluminum, cadmium,
calcium, chromium, copper, iron, lead, magnesium, molybdenum, nickel, and
zinc have been grouped for ease of presentation.  Arsenic, mercury, and
selenium are treated separately.
*                                    .
 References can be found on page 3-136.
                                 3-77

-------
                       Sample Handling and Storage

             A generalized flowchart for the processing of water and
sediment samples to "be analyzed for metals is presented in Figure 3-8.
The chart covers both water and sediment samples and all three chemical
tests discussed earlier in the manual.  The intent is to reemphasize
that different methods of sample handling and pretreatment are required
for each test.  The selection of a specific test (bulk analysis,
elutriate test, etc.) depends on the purpose of the .study.
             Water samples may be split into three fractions.  The first
(Wl), consisting of the unfiltered water, can be digested as discussed
later to provide a measure of the total metal concentration in the sample.
The second fraction (W2) is a filtered water sample to be analyzed for
soluble metal concentrations.  The third fraction (W3) is an unfiltered,
unpreserved water sample to be used in the elutriate test.  Recommenda-
tions for sample pretreatment, preservation, and storage time are
presented in Figure 3-8.
             Sediment samples may be handled in three different ways
although the selection of a storage method may limit the future use of
the sample.  For example, a sediment sample stored in a moist condition
at 1*C (Si) can be used in the elutriate test, element partitioning
studies, bioassays, or total analysis.  However, sediment samples stored
in a dried (S2) or frozen (S3) state should only be used for total or
bulk analysis.  Methods are presented for the digestion of sediment
samples prior to analysis.  The most frequently used digestion methods
utilize a combination of hydrochloric acid and nitric acid since more
severe treatment with hydrofluoric acid or perchloric acid requires
special equipment or hoods, and other methods are less reliable or
reproducible.6
             There are other available methods such as neutron activation
analysis that are specific and more sensitive than atomic absorption.
However, the present limited availability of the necessary instrumentation
does not justify inclusion in this version of this manual.
                                  3-78

-------

CORE SAMPLE
4

'
WATER SAMPLE DREDGE SAMPLE CORE SECTION
4 4
444 * i
'
'
1 ACIDIFY FILTER NO TREATMENT STORE WET DRY
1 	 ,
.
STORE ACIDIFY STORE
i
DIGEST STORE
\ '
4 * ' 4 4 i

^
FREEZE
1
STORE
1
_..to FIMTRIiTE FRAr.TinNATF RIOASSAY DIGEST DIGEST
** (SIC)
i

1
1 ANALYZE ANALYZE ANALYZE ANALYZE ANALYZE ANALYZE
| (Wl) (W2) (S1A) (SIB) (SID) (S2)
U>
^ SAMPLE DESIGNATION
PURPOSE
CONTAINER
SAMPLE TREATMENT
PRESERVATIVE
Wl W2 W3 S1A SIB SIC S10
Total Water Soluble Used in Mobile Chemical Bioavall- Total
Cone. Water Elutriate Cone. Distribution ability Sediment
Cone. Cone.
G,P G,P G, P G, P G, P G. P G, P
None Filter None None None None None
HN03 HN03 None ^"C ^"C 
-------
                      Procedures for Water Samples

                   (All Metals Except As, Hg, and Se)


Method 1:  Direct Flame Atomic Absorption, Total Metals (Wl)5

Apparatus

Atomic absorption spectrophotometer:  for samples vith a high salt content,
      a deuterium background corrector or a double-beam instrument would
      be desirable

Burner:  the most common type of burner, known as a premix, introduces the
      spray into a condensing chamber for removal of large droplets.
      The burner may be fitted with a conventional head containing a
      single 3-in.-long (7-6 cm) slot for aspirating organic solvents,
      a three-slot Boling head for direct aspiration of aqueous samples
      into an air-acetylene flame, or a head containing a single 2-in.
      (5 cm) slot for use with a nitrous oxide-acetylene flame

Recorder

Hollow cathode lamps:  multielement lamps are available but not
      recommended.  A separate lamp should be used for each metal to
      be determined
Reagents

Air:  cleaned and dried through a suitable filter to remove oil, water,
      and other foreign substances.  The source may be a compressor or
      commercially bottled gas.
Acetylene:  standard commercial grade.  Acetone, which is always present
      in acetylene cylinders, can be prevented from entering and damaging
      the burner head by replacing a cylinder when its pressure has
      fallen to 7 kg/cm  (100 psig) acetylene.
Nitrous oxide:  commercially available cylinders.
Calcium solution:  dissolve 630 mg calcium carbonate, CaCOa, in 10 ml
      concentrated HC1.  Add 200 ml water and, if necessary, heat the
      solution and boil gently to obtain complete solution.  Cool and
      dilute to 1000 ml with deionized distilled water.

Deionized distilled water:  use deionized distilled water for the
      preparation of all reagents and calibration standards and as
      dilution water.
Hydrochloric acid:  HC1, concentrated.

Lanthanum solution:  dissolve 58.65 g lanthanum oxide, LaaOa, in 250 ml
      concentrated HC1.  Add the acid slowly until the material is
      dissolved and dilute to 1000 ml with deionized distilled water.
Nitric acid:  HNOs, concentrated.
                                  3-80

-------
Sodium chloride solution:  dissolve 250 g NaCI in deionized distilled
      water and dilute to 1000 ml.

Standard metal solutions:  prepare a series of standard metal solutions
      containing 5 to 1000 ug/ by appropriate dilution of the following
      stock metal solutions with deionized distilled water containing
      1.5 ml concentrated HNOa/C.

Aluminum:  dissolve 1.000 g aluminum metal in 20 ml cone. HC1 by heating
      gently and diluting to 1000 ml, or dissolve 17.581* g aluminum
      potassium sulfate (also called potassium alum), AlK(SOit )z    12 IfeO,
      in 200 ml deionized distilled water, add 1.5 ml cone. HNOa,  and
      dilute to 1000 ml with deionized distilled water; 1.00 ml = 1.00 mg
      Al.
Calcium:  to 2.^972 g calcium carbonate, CaC03 , add 50 ml deionized water
      and add dropwise a minimum volume of cone. HC1 (about 10 ml) to
      effect complete solution.  Dilute to 1000 ml with deionized
      distilled water; 1.00 ml = 1.00 mg Ca.
Cadmium:  dissolve 1.000 g cadmium metal in a minimum volume of 1+1 HC1.
      Dilute to 1000 ml with deionized distilled water; 1.00 ml = 1.00 mg
      Cd.
Chromium:  dissolve 2.828 g anhydrous potassium dichromate, K^C^O?, in
      about 200 ml deionized distilled water, add 1.5 ml cone. HNOa, and
      dilute to 1000 ml with deionized distilled water; 1.00 ml = 1.00 mg
      Cr.
Copper:  dissolve 1.000 g iron wire in 50 ml of 1+1 HNOs and dilute to
      1000 ml with deionized distilled water; 1.00 ml = 1.00 mg Cu.

Iron:  dissolve 1.000 g iron wire in 50 ml of 1+1 HNOs and dilute to
      1000 ml with deionized distilled water; 1.00 ml = 1.00 mg Fe.

Lead:  dissolve 1.598 lead nitrate, Pb(N03)2, in about 200 ml of water,
      add 1.5 ml cone. HNOs, and dilute to 1000 ml with deionized
      distilled water; 1.00 ml = 1.00 mg Pb.

Magnesium:  dissolve 10.0135 g magnesium sulfate heptahydrate , MgSOit 
      7H20, in 200 ml deionized distilled water, add 1.5 ml cone. HN03,
      and, make up to 1000 ml with deionized distilled water; 1.00 ml =
      1.00 mg Mg.

Manganese:  dissolve 3.076 g manganous sulfate monohydrate, MnSOi* 
      HaO, in about 200 ml deionized distilled water, add 1.5 nil cone.
      HNOa, and make up to 1000 ml with deionized distilled water;
      1.00 ml = 1.00 mg Mn.

Molybdenum:  dissolve 1.8UO g ammonium molybdate, (NHiOetMoyOz) '
      UHaO, in deionized water and dilute to 1 A.

Nickel:  dissolve U.953 g nickelous nitrate hexahydrate, Ni(N03)2 '
      6H20, in about 200 ml deionized distilled water, add 1.5 ml cone.
      HN03, and make up to 1000 ml with deionized distilled water;
      1.00 ml = 1.00 mg Ni.
                                 3-81

-------
Zinc:  dissolve 1.000 g zinc metal in 20 ml 1+1 HC1 and dilute to 1000 ml
      with deionized distilled water; 1.00 ml = 1.00 mg Zn.
             Prepare dilute working solutions of each metal as required.
These solutions should be prepared fresh as needed.  In addition, the
following steps are necessary for the preparation of working solutions
of calcium, magnesium, iron, and manganese.  Calcium and magnesium
analyses are subject to pH effects and to interference from aluminum
when analyzed by atomic absorption.  These effects can be overcome
by mixing 100 ml calcium and magnesium standard with 25 ml lanthanum
chloride solution prior to aspiration.5  Atomic absorption analysis
for iron and manganese is also subject to an interference that can be
overcome by mixing 100 ml working standard solution with 25 ml of
calcium chloride solution prior to aspiration.
Sample preparation
             Transfer a 50- to 100-ml, well-mixed 1 sample to a 150-ml
or larger beaker.  Add 5 ml concentrated HNOs to the sample and evaporate
to near dryness on a hot plate.  Caution should be exercised during this
process to ensure that the sample does not boil.
             Cool the sample and add a second 5-ml portion of concen-
trated HNOs,  Cover the beaker with a watch glass and reflux the sample
on a hot plate.  Additional acid should be added to the sample as
necessary during the refluxing.  The heating should be continued until
the digestion process is complete, as indicated by the presence of a
light-colored residue.
             Following digestion, add 1 to 2 ml concentrated HC1 and
warm the beaker slightly.  Wash the watch glass and beaker walls with
distilled water.  Filter the digestate to remove any remaining insoluble
matter and adjust the volume of the filtrate to a convenient volume with
distilled water.
             Analyze the sample and report results as total concentration.
Quantification procedure
             Prepare a series of working metal standards by diluting the
appropriate stock solutions with deionized distilled water containing
1.5 ml concentrated HN03/&.  These solutions should be prepared fresh
on the day of use.
                                  3-82

-------
             Install the appropriate hollow cathode lamp in the instru-
ment.  Align the lamp and set the source current according to the manu-
facturer's instructions.  Turn on the instrument and allow "both the
instrument and lamp to warm up.  This process usually requires 10 to
20 min.
             Set the wavelength dial according to Table 3-9    The infor-
mation in Table 3-9 should only be used as a guide in setting up the
instrument.  Due to calibration differences, the actual wavelength should
be based on maximum sensitivity after the instrument has completely
warmed up.  Set the slit width according to manufacturer's instructions.
             Install the burner head indicated in Table 3-9-
             Turn on appropriate gases, ignite flame, and adjust the
flow of fuel and oxidant to give maximum sensitivity for the metal being
measured.  When using a nitrous oxide flame, a T-junction valve or
alternate switching valve should be employed for rapidly changing from
nitrous oxide to air to prevent flashbacks when the flame is turned on
or off.
             Atomize deionized distilled water acidified with 1.5 ml
concentrated HNO /$, and check the aspiration rate for 1 min.   If
necessary, adjust the aspiration rate to 3 to 5 ml/min.  Zero the
instrument.
             Atomize a standard and adjust the burner alignment (up,
down, sideways) until a maximum signal response is obtained.
             Aspirate a series of metal standards that bracket the
expected range of sample concentrations and record the absorbance of
each standard.  Rinse the atomizer with deionized distilled water con-
taining 1.5 ml concentrated HNOs/Jl between each standard.
             Atomize the digested water samples (Wl) and determine their
absorbances.  Rinse the atomizer with dilute nitric acid between each
sample.
             Samples scheduled for iron and manganese or calcium and
magnesium analysis should be premixed with calcium chloride or lanthanum
chloride solution, respectively.  This is accomplished by mixing four
volumes of digested sample with one volume of the appropriate salt

                                    3-83

-------
solution.
             When determining metal concentrations "by atomic absorption,
the following sequence of sample processing is recommended:
             a_.  Run a set of standards.
             b_.  Run five samples.
             c_.  Run a duplicate of the fifth sample.
             d_.  Run five additional samples.
             e_.  Run a duplicate of the fifth sample.
             f_.  Run a fifth sample that has been spiked.
             g_.  Run a standard.
             h,.  Repeat Steps _a through .
             i_.  Rerun standards.
This approach will incorporate certain aspects of the quality control
program into the analytical procedure.  Specifically, the suggested
sequence allows for evaluation of instrument stability, replicate
analysis, and spike recovery.
Calculations
             Prepare a standard curve by plotting the absorbance of each
standard versus concentration for each metal.  Use the standard curve
to convert sample absorbance to metal concentration.
                                 3-8U

-------
Method 2:  Direct Flame Atomic Absorption, Soluble Metals (W2,  S1A)5
Apparatus
Atomic absorption spectrophotometer and appropriate burner head(s)
Recorder
Hollow cathode lamps
Reagents
Air
Acetylene
Nitrous oxide
Deionized distilled water
Concentrated hydrochloric acid
Concentrated nitric acid
Stock metal solutions as described earlier
Sample preparation
             The operational definition of a soluble metal concentration
is one that passes a membrane filter, usually of O.h^-v pore-size diameter.
Thus, both W2 and S1A are filtered samples.  The W2 samples should be
filtered at the time of collection, if possible, or as soon as  practical
thereafter.  The S1A samples are filtered as part of the preparation  of
the standard elutriate.
             Both of these samples may be analyzed without further
treatment and the results should be reported as soluble or filtrable.
Quantification procedure
             Prepare a series of working metal standards by diluting  the
appropriate stock solutions with deionized distilled water containing
1.5 ml concentrated HNOa/Jc.  These solutions should be prepared fresh
on the day of use.
             To minimize possible matrix effects, samples should also
contain 1.5 nil concentrated HNOa/Jl.  W2 samples should routinely be
preserved with HNOs at the time of filtration, but it will be necessary
to add the appropriate volume of HNOa to the S1A samples.
             Install the appropriate hollow cathode lamp in the instru-
ment.  Align the lamp and set the source current according to the
manufacturer's instructions.  Turn on the instrument and allow  both the
                                 3-85

-------
instrument and the lamp to warm up.   This process usually requires 10 to
20 min.
             Set the wavelength dial according to Table 3-9-  The infor-
mation in Table 3-9 should only be used as a guide in setting up the
instrument.  Due to calibration differences, the actual wavelength
should be based on maximum sensitivity after the instrument has com-
pletely warmed up.  Set the slit width according to manufacturer's
instructions.
             Install the burner head indicated in Table 3-9-
             Turn on appropriate gases, ignite flame, and adjust the
flow of fuel and oxidant to give maximum sensitivity for the metal being
measured.  When using a nitrous oxide flame, a T-junction valve or
alternate switching valve should be employed for rapidly changing from
nitrous oxide to air to prevent flashbacks when the flame is turned on
or off.
             Atomize deionized distilled water acidified with 1.5 nil
concentrated HNOs/& and check the aspiration rate for 1 min.  If neces-
sary, adjust the aspiration rate to 3 to 5 ml/min.  Zero the instrument.
             Atomize a standard and adjust the burner alignment (up,
down, sideways) until a maximum signal response is obtained.
             Aspirate a series of metal standards that bracket the
expected range of sample concentrations and record the absorbance of
each standard.  Rinse the atomizer with deionized distilled water
containing 1.5 ml concentrated HNOs/X, between each standard.
             Atomize the water samples (W2, S1A) and determine their
absorbances.  Rinse the atomizer with dilute nitric acid between each
sample.
             Samples scheduled for iron and manganese or calcium and
magnesium analysis should be premixed with calcium chloride or lanthanum
chloride reagents, respectively, as discussed earlier.  This is accom-
plished by mixing four volumes of filtered water sample with one volume
of the appropriate reagent.
             When determining metal concentrations by atomic absorption,
                                 3-86

-------
                   Table 3-9


Recommended Atomic Absorption Spectrophotometer

                                         7 8
   Instrument Settings for Metal Analysis '

Metal
Al
As
Cd
Ca
Cr
Cu
Fe
Pb
Mg
Mn
Hg
Mo
Ni
Se
Zn
Slit
)|

Ij.
k
k
k
3
*
k
h
6
k
3

k
Wavelength.
mu
309
193
228
U22
357
32U
2U8
283
285
279
253
313
232
196
213
.3
.7
.8
.7
.9
.7
.3
.3
.2
.8
.7
.5
.0
.0
.9
Burner
Nitrous oxide

Boling
2-slot
sideways
3-slot

Boling
Boling
Nitrous oxide
sideways
Boling

Nitrous oxide
Boling

3-slot
Fuel
Acetylene

Acetylene
Acetylene
Acetylene
Acetylene
Acetylene
Acetylene
Acetylene
Acetylene

Acetylene
Acetylene

Acetylene
Oxidant
Nitrous

Air
Air
Air
Air
Air
Air
Nitrous
Air

Nitrous
Air

Air
oxide







oxide


oxide



                     3-87

-------
    following sequence of sample processing is recommended:
             a_.  Run a set of standards.
             b_.  Run five samples.
             c_.  Run a duplicate of the fifth sample.
             d_.  Run five additional samples.
             __.  Run a duplicate of the fifth sample.
             f_.  Run a fifth sample that has been spiked.
             g_.  Run a standard.
             h..  Repeat Steps a_ through f_.
             i_.  Rerun standards.
In this way, the time stability of the instrument can be checked and an
analytical quality control program can be incorporated into the sample
processing routine.
Calculations
             Prepare a standard curve by plotting the absorbance of each
standard versus concentration for each metal.  Use the standard curve to
convert sample absorbance to metal concentration.

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Method 3:  Graphite Furnace Atomic Absorption5'8
             The use of graphite furnaces or carbon rod atomizers is
considered to be an approved test method because it is essentially an
atomic absorption technique.9  However, the method is not meant for use
with all samples.  The method should only be considered as an alternative
to conventional flame atomic absorption spectrophotometry when one or
more of the following conditions exist:10
             a_.  Greater analytical sensitivity is required.
             b_.  Sample size is limited.
             c_.  Samples have a high dissolved solids content and cannot
                 be aspirated into a flame.
These conditions will usually result in the use of a graphite furnace
with water samples because of the higher metal concentrations in sediments
and the relative ease of increasing sediment sample size during the
digestion step.
Apparatus
Atomic absorption spectrophotometer equipped with a deuterium background
      corrector.  A double beam instrument would be considered preferable
Graphite furnace or carbon rod attachment
Automatic sampler attachment for the atomic absorption spectrophotometer
      or an Eppendorf pipette
Recorder
Hollow cathode lamps, a separate lamp for each metal to be determined
NOTE:  The automatic sampler is the preferred method of sample transfer
       because it reduces analytical variability compared to  other
       methods of sample pipetting.
Reagents
Reagents are the same as those used for conventional flame atomic
      absorption spectrophotometry.
Sample preparation
             Graphite furnace analyses require small sample volumes.
In order to avoid the problem of obtaining representative subsamples
from heterogeneous samples when using small sample sizes, samples
should be pretreated as discussed with Method 1.   Soluble metal
concentrations can be determined directly on filtered water samples
(W2, S1A).  Total metal concentrations can be determined with the
                                 3-89

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graphite furnace technique following strong acid digestion of the
samples.  Sediment digests (.SID, S2, S3)  can also be analyzed with the
aid of a graphite furnace "but consideration should be given to digesting
a larger sediment sample to achieve the desired increase in sensitivity.
The digest can then be analyzed with conventional flame atomic absorption
spectrophotometry.
Quantification procedure
             Once a decision has been made to utilize a graphite
furnace, install the attachment according to manufacturer's instruc-
tions.  Align the atomizer as required and warm up the instrument.
Optimum instrument conditions for the graphite furnace will generally
be identical to those used for flame atomization.  Check individual
operation manuals for differences with specific metals (usually arsenic
and selenium).  Warm up the background corrector, which should always
be used with the graphite furnace.
             Prior to the analysis of a new series of samples or the
use of a new sample cup, it is recommended that the atomizer be decon-
taminated.  This can be accomplished by operating the instrument in the
maximum temperature mode for approximately 2 sec.
             The analysis is conducted by transferring a known volume
of standard or sample, 5 to 20 yi, to the sample cup.   The recommended
method of sample transfer would be through the use of an automatic
sampler attachment.  Other methods such as the use of oxford or
Eppendorf pipettors can be used; but an increase in analytical
variability is to be expected as a result.  The sample is then sub-
jected to a drying cycle to remove solvent, an ashing cycle to destroy
organic matter, and an atomizing cycle (Table 3-10)..   The metal
concentration is quantified during the atomization cycle.  The selection
of drying temperature and time, ashing temperature and time, and
atomization temperature and time is  critical if reproducible  results
are to be obtained.    Manufacturer's instructions should be followed.
             Process samples in the same order suggested for conven-
tional flame analysis:  run a series of known standards to prepare a
calibration curve; run five samples and duplicate the fifth-sample;
                                 3-90

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                                                   Table 3-10

                            Graphite Furnace Operating Conditions for Selected Metals
          Conditions       Al     As     Cd     Cr     Cu    Fe     Fb     Mn     Mo     Ni     Se     Zn
       Drying Time,
        sec                30     30     30     30     30     30     30     30     30      30      30      30


       Drying Temp.,
       C               125    125    125    125    125    125    125    125    125     125     125     125


       Ashing Time,
        sec                30     30     30     30     30     30     30     30     30      30      30      30
u>
VQ
H      Ashing Temp.,
        C              1300   1100    500   1000    900   1000    500   1000   1^00     900   1200    UOO


       Atomizing Time,
        sec
10     10     10     10     10     10     10     10     15     10     10     10
       Atomizing Temp.,
        C               2700    2700   1900   2700   2700   2700   2700   2700   2800   2700   2700   2500


       Purge Gas         ArArArArArArArArArArArAr



       Wavelength,
        nm                309.3  193.7   228.8  357.9  32^.7  2U8.3  283.3  279-5  313.3  232.0  196.0  213.9

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run five more samples;  run a duplicate and a known  spike; run a standard
to check instrument stability;  and repeat the cycle.
Calculations
             Calculate sample concentrations by using  a calibration
curve to convert sample absorbance to concentration.
                                  3-92

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Method h:  Chelation-Extraction Atomic Absorption8
             This method of quantifying metals is another  version of
atomic absorption spectrophotometry.   The advantage of this method is
the increased sensitivity that is obtained by chelating metals  with an
organic ligand and concentrating the  metal complexes into  an  organic
solvent.  This approach offers the advantages of concentrating  the
sample (.by the ratio of the initial sample volume to the volume of
organic solvent) and removing the metal from any interfering  substances
in the original sample.  However, the sample pH can affect extraction
efficiency7'10 and the extracted metal complex is time dependent.
Thus, the extracts should be processed promptly. In addition,  the
metals must be analyzed in a specific order with the least stable metal
complexes being analyzed first and the most stable  metal complexes being
analyzed last.
Apparatus
pH meter
Shaker, mechanical, and holders for 250-ml volumetric flasks
Glassware:  all glassware such as Erlenmeyer flasks, beakers, pipettes,
      volumetric flasks, and funnels  should be rinsed with 1:1  nitric
      acid and rinsed with deionized  water
Atomic absorption spectrophotometer,  equipped with  appropriate  burner(s)
Recorder
Reagents
Ammonium pyrrolidine dithiocarbamate  (APDC) solution, 1 percent:
      dissolve 1 g APDC in 100 ml deionized water and filter, if
      necessary.  Prepare fresh before use.
Methyl isobutyl ketone (MIBK).
Buffer (pH U.T5) solution:  dissolve  272 g sodium acetate, CHaCOOKa, in
      distilled water and dilute to about 1  in a  2- beaker.   Add
      acetic acid, CH3COOH, to the solution until a pH of  U.T5  is
      reached (use a pH meter).  Dilute to 2 &. Extract this solution
      with 0.01 percent dithizone until the extract remains
      green; then extract with carbon tetrachloride, CC1 , to remove
      excess dithizone.
Dithizone, 0.01 percent:  dissolve 0.01 g of diphenylthiocarbazone in
      100 ml carbon tetrachloride.
Stock metals solutions, prepare as described earlier.
                                 3-93

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Sample preparation
             Pipette a 100-ml volume of sample digest  (Wl,  W2,  S1A)
into a 250-ml volumetric flask.   If more sensitivity is desired,  a
larger sample size can be used.   However, the size of the flask and
the volume of subsequent reagents should be increased  proportionately.
             Prepare a blank and sufficient standards  to cover  the
expected concentration range of the samples.
             Add 5-0 ml buffer to each and mix by swirling flasks.
Adjust the pH in the range of 2 to 3 using either bromphenol  blue or
a pH meter.
             Add 5-0 ml APDC to each and mix  by swirling flasks.
             Add 10.0 ml MIBK and shake the flasks for 5 min  on a
mechanical shaker equipped with special holders for the flasks.
             Allow the solvent layers to separate.  In order  to raise
the MIBK layer to the neck of the flask, slowly add deionized water
down the side of the flask.  This will reduce the number of sample
transfers and the amount of glassware required.  However, it  is
essential that the water be added very carefully as physical  distur-
bance of the organic layer could reduce the extraction efficiency.
             It is imperative to point out that the stability of  the
extracted APDC-metal complexes is time dependent and generally  on the
order of a few hours.  Therefore, it is necessary that sample analysis
commence immediately after extraction.
Quantification procedure
             Install the appropriate hollow cathode lamp in the
instrument.  Align the lamps and set the source current according to
the manufacturer's instructions.  Turn on the instrument and  allow
both the instrument and lamps to warm up.  This process usually
requires 10 to 20 min.  If possible, this process should be started
before the extractions are completed because  of the limited stability
of the metal extracts.
             Install a single slot burner designed for use with
organic solvents.  Turn on the appropriate fuel and oxidant gases and
adjust the flows to give maximum sensitivity  for the metal being

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measured.  In setting the fuel-to-air ratio of the gas mixture at  the
turner, start with the settings recommended by the manufacturer for
the analysis (Table 3-9)   Note the color of the flame and begin
aspiration of MIBK solvent.  Gradually reduce the flow of the fuel
while continuing to aspirate the solvent until the color of the flame
is similar to that noted earlier.  This step is necessary because  the
organic solvent contributes to the fuel supply.  If this precaution  is
not taken, the resultant flame may produce an undesirable luminescence
or be lifted off the burner.  With the nitrous oxide burner, approxi-
mately double the flow of acetylene is required over that normally used
for the air acetylene burner; the acetylene should be adjusted until the
flame is a rose-red color.
NOTE:  When using a nitrous oxide flame, a T-junction valve or alternate
       switching valve should be employed for rapidly changing from
       nitrous oxide to air to prevent flashbacks when the flame is
       turned on or off.
             Atomize the organic solvent and check the aspiration  rate
for 1 min.  If necessary,  adjust the aspiration rate to 3 to 5 ml/min.
Zero the instrument.  Atomize a standard and adjust the burner align-
ment until a maximum signal response is obtained.
             Aspirate the extracted metal standards and record the
absorbance of each standard.'  Rinse the burner with organic solvent
between each standard.
             Aspirate the solvent layer from each of the extracted
samples.  Record the sample absorbance.  Continue to rinse the burner
with organic solvent between each sample.
             The sequence of sample processing should consist of a set
of standards, five samples, a duplicate sample, a standard, five
samples, a duplicate sample, and a 'spiked sample.  Repeat the sequence
until all samples have been processed.
Calculations
             Prepare a calibration curve derived from the peak heights
obtained with the standard solutions.  Determine the concentration of
metal in the sample by comparing sample peak height with the calibration
curve.
                                3-95

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                       Procedure for Sediment Samples
                     (All Metals Except As, Hg, and Se)

Method 1:  Direct Flame Atomic Absorption, Total Metals
           (SID, S2, S3)5'11
Apparatus
Atomic absorption spectrophotometer with appropriate "burner head(s)
Recorder
Hollov cathode lamps
Balance
Digestion apparatus (hot plate, Kjeldahl digestion unit,  muffle
      furnace, or digestion bomb)
Reagents
Air
Acetylene
Nitrous oxide
Deionized distilled water
Stock metal solutions, as described earlier
Digestion reagents, as specified with each digestion procedure
Sample preparation
             Each of the designated fractions, SID, S2, and S3,  listed
with this procedure is intended for strong acid digestion and total
metal analysis.  The only difference between these samples is the
method of handling prior to digestion:   one sample is stored moist and
has a shorter recommended storage period (SID); one sample has been
previously air dried (S2); and the third sample has been  stored
frozen (.S3).   The selection of a sample storage method is left up to
each laboratory although the use of moist samples (SID) is recommended
because the same sediment could be used in the elutriate  test,
elemental partitioning, and bioassay studies, if necessary.12
             Depending on the method of sample storage, proceed with
the appropriate procedure:
             a_.  Starting with, moist sediment samples (SID), determine
                 the percent solids in the samples.  Accurately weigh
                 a 0.5- to 1.0-g dry weight equivalent aliquot of the
                                 3-96

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                 sample using an analytical "balance.   The sample weight
                 should "be selected based on the anticipated metal
                 concentrations and the detection limit and/or the
                 upper concentration range of the analytical technique
                 to "be used.
             b_.  Starting with dried sediment samples (S2),  accurately
                 weigh a 0.5- to 1.0-g aliquot of the sample using an
                 analytical balance.  The sample weight should be
                 selected based on the anticipated metal concentrations
                 and the detection limit and/or the upper concentration
                 range of the analytical technique to be used.
             c_.  Starting with a frozen sediment sample (S3), thaw the
                 samples at room temperature.  Determine the percent
                 solids in the samples.  Accurately weigh a  0.5- to
                 1.0-g dry weight equivalent aliquot of the  sample using
                 an analytical balance.  The sample weight should be
                 selected based on the anticipated metal concentrations
                 and the detection limit and/or the upper concentration
                 range of the analytical technique to be used.
             Proceed with one of the digestion techniques provided in
Tables 3-11 through 3-17.  Treatment with hydrofluoric acid  and
perchloric acid (Table 3-l6")  or treatment with hydrofluoric  acid-nitric
acid-hydrochloric acid (.Table 3-17) are the most severe of the listed
procedures and regarded as yielding the most complete digestion.  How-
ever, these procedures should only be used with appropriate  hoods and
safety equipment.  Combinations of nitric acid and hydrochloric acid
(Table 3-11) have been proven to be effective digestion solutions for
routine use and do not require special hoods or glassware.
             The procedures summarized in Tables 3-11 through 3-15
were evaluated to determine their ability to solubilize 13 metals
(.As, Cd, Cr, Cu, Pb, Ni, Ca,  Mg, Na, K, Fe, Mn, and Zn) from  a municipal
treatment plant sludge.6  Based on calculated 95 percent confidence
limits, the HNOs-HCl digestion procedure in Table 3-10 provided
equivalent recoveries for all metals.  The HNOs digestion procedure
(Table 3-12) was comparable to the HNOs-HCl digestion procedure for
all metals except Ni.  Low As and Ca results were observed with HN03-
HaOa treatment (Table 3-13).   The least satisfactory results were
observed with muffle furnace ignition (Table 3-lJO and low temperature
ashing (Table 3-15).  Muffle furnace ignition produced low results for
                                3-97

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                                 Table 3-11
                           M03-HC1 Digestion6'13
                                      Procedure
 1.      Prepare HN03-HC1 (l:3 v/v) digestion mixture (aqua regia) just
         prior to each use by carefully adding, with stirring, one volume
         cone. HNO to three volumes of cone. HC1.   A convenient batch vol-
         ume is a 30 ml cone. HN03 and 90 ml cone.  HC1.
         CAUTION:  Avoid inhaling fumes.

 2.      Accurately weigh a 0.05- to 1.0-g dried sludge sample, using
         an analytical balance.  Select the sample weight based on antici-
         pated metal concentrations and the detection limit/upper concen-
         tration range of the atomic absorption spectrophotometer calibra-
         tion curves.

 3.      Place dried sludge in an Erlenmeyer flask (125 or 250 ml volume).
         Alternatively, a 250-ml beaker with watch glass may be used.

 k.      Moisten dried sludge with ca. 0.5 to 1.0 ml deionized distilled
         water (DDW).

 5.      Slowly add 10 ml HN03-HC1 and swirl container to control effer-
         vescence and to ensure good mixing.

 6.      Place container on hot plate.  Bring to slow boil.   Continue boil-
         ing until solution approaches dryness.

 T.      Carefully add more HN03-HC1 in 5-ml increments and repeat Step 5
         until all visible organic matter is destroyed and the solution
         begins to clear.

 8.      Continue boiling until the evolution of reddish-brown fumes
         ceases.

 9.      Remove container from hot plate, cool to room temperature, add ca.
         20 ml DDW, and separate the digestate from any mineral residue, if
         present, by filtering through a Whatman No.  1*2 or equivalent  filter
         paper or a O.i|-ym membrane filter.
10.      Rinse container and filter paper with ca.  5 to 10 ml DDW two  times
         and collect rinses.  Quantitatively transfer and combine rinses
         and filtrate into a volumetric flask (50 or 100 ml volume) and
         dilute with DDW to the volume mark.
                                (Continued)
                                     3-98

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                         Table 3-11 (Concluded)
                                      Procedure
11.      Dilute the solution from Step 10 further,  if necessary.   Analyze
         metal(s) by atomic absorption spectrophotometry according to the
         instrument manufacturer's operating instructions.   Calculate and
         report the concentration of metal(s)  in the sludge sample on a
         mg/kg dry weight basis.
                                  3-99

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                             Table 3-12
                           HW03 Digestion6
                                  Procedure
1.      Place dried sludge in an Erlenmeyer  flask (125  or  250 ml
        volume).   Alternatively, a 250-ml beaker  with watch glass may
        be used.
2.      Moisten dried sludge with ca.  0.5 to 1.0  ml deionized distilled
        water (DDW).
3.      Slowly add 10 ml cone.  HNOs  and swirl container to control
        effervescence and to ensure  good mixing.
k.      Place container on hot plate.   Bring to slow boil.   Continue
        boiling until the solution approaches dryness.
5.      Carefully add more cone. HNOa  in 5-ml increments and repeat
        Step 5 until all visible organic matter is destroyed and the
        solution begins to clear.
6.      Continue boiling until the evolution of reddish-brown fumes
        ceases.
7.      Remove container from hot plate, cool to  room temperature, add
        ca.  20 ml DDW, and separate  the digestate from  any mineral
        residue,  if present, by filtering through a Whatman No. h2 or
        equivalent filter paper or a O.U-p membrane filter.   Collect
        filtrate.
8.      Rinse container and filter paper with ca.  5 to  10  ml DDW two
        times and collect rinses.   Quantitatively transfer and combine
        rinses and filtrate into a volumetric flask (50 or 100 ml
        volume) and dilute with DDW to the volume mark.
9.      Dilute the solution from Step  9 further,  if necessary.  Analyze
        metal(s)  by atomic absorption  spectrophotometry according to
        the instrument manufacturer's  operating instructions. Calculate
        and report the concentration of metal(s)  in the sludge sample
        on a mg/kg dry weight basis.

                               3-100

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                               Table 3-13
                          HNOa-H202  Digestion6
Step                                Procedure
 1.      Place dried sludge in an Erlenmeyer flask (125 or  250 ml  volume)
         Alternatively, a 250-ml beaker with watch glass may  be used.
 2.      Moisten dried sludge with ca.  0.5 to 1.0 ml deionized distilled
         water (DDW).
 3.      Slowly add 10 ml cone.  HNOa  and swirl container to control
         effervescence and to ensure good mixing.
 k.      Place container on hot plate.   Bring to slow boil.   Continue
         boiling until the solution approaches dryness.
 5.      Carefully add more cone. HN03  in 5-ml increments and repeat
         Step h until all visible organic matter is destroyed and  the
         solution begins to clear.
 6.      Continue boiling until the evolution of reddish-brown fumes
         ceases.
 T.      Remove container from the hot  plate and cool to ca.  room
         temperature.
 8.      Add 3 ml cone. HN03 and 10 ml  30 percent H202. Return con-
         tainer to the hot plate and warm gently.   Using tongs,
         alternatively remove the container from the hot plate to  allow
         any effervescence to subside and then rewarm.   Continue this
         process until subsequent warming does not produce  any further
         effervescence.
 9.      Heat the solution to boiling and continue heating  for at  least
         15 min.
10.      Remove container from hot plate, cool to room temperature, add
         ca. 20 ml DDW, and separate the digestate from any mineral
         residue, if present, by filtering through a Whatman  No. 1*2 or
         equivalent filter paper or a 0.^5-y membrane filter.  Collect
                               (Continued)
                                  3-101

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                         Table 3-13 (Concluded)
                                    Procedure
10.      filtrate.
11.      Rinse container and filter  paper  with  ca.  5 to  10 ml DDW two
         times and collect rinses.   Quantitatively  transfer and combine
         rinses and filtrate into  a  volumetric  flask (50 or 100 ml
         volume) and dilute with DDW to the  volume  mark.
12.      Dilute the solution from  Step  11  further,  if  necessary.
         Analyze metal(s)  by atomic  absorption  spectrophotometry accord-
         ing  to the instrument manufacturer's operating  instructions.
         Calculate and report the  concentration of  metal(s) in the
         sludge sample on a mg/kg  dry weight basis.
                                3-102

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                               Table 3-lh
                        Muffle Furnace Ignition6
Step                                Procedure
 1.      Place the Pt crucible containing the dried sludge  sample into
         a room temperature muffle furnace.  Bring the furnace to 550C
         and maintain the temperature  for ca. 30 min.
 2.      Remove the Pt crucible from the muffle furnace, cool for 5 to
         10 min, place in a desiccator, and  cool to room temperature.
         Reweigh the crucible, if desired, to determine the amount of
         volatile matter lost on ignition.
 3.      Add a small volume (l to 3 ml) of warm cone. HNOs  to the residue
         in the Pt crucible and place  on a hot plate.  Heat the crucible
         (avoid splattering and do not boil) until most of  the acid has
         evaporated.  Do not heat to dryness.
 U.      Remove container from hot plate, cool to room temperature, add
         ca. 20 ml DDW, and separate the digestate from any mineral
         residue, if present, by filtering through a Whatman No. k2 or
         equivalent filter paper or a  O.^-y  membrane filter.  Collect
         filtrate.
 5.      Rinse container and filter paper with ca. 5 to 10  ml DDW two
         times and collect rinses.  Quantitatively transfer and combine
         rinses and filtrate into a volumetric flask (50 or 100 ml
         volume) and dilute with DDW to the  volume mark.
 6.      Dilute the solution from Step 5 further, if necessary.  Analyze
         metal(s) by atomic absorption spectrophotometry according to
         the instrument manufacturer's operating instructions.  Calculate
         and report the concentration  of metal(s) in the sludge sample
         on a mg/kg dry weight basis.
                                 3-103

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                              Table 3-15
                        Low Temperature Ashing6
 ep                                Procedure
1.      Place the Petri dish containing the  dried  sludge  sample  into the
        low temperature ashing instrument cavity and  operate according
        to the instruction manual.   A flat quartz  plate can "be installed
        in cylindrical cavities for ease of  sample placement.  Experi-
        ence showed that three successive 8-hr  ashing cycles at  200 watts
        (RF) were necessary to completely ash the  sludge  sample.
2.      Upon completion of the ashing step,  dissolve  the  residue with a
        small volume (ca.  1 to 3 ml)  of cone. HNOa and quantitatively
        transfer the solution to a  filtration apparatus,  if necessary.
3.      Rinse container and filter  paper with ca.  5 to 10 ml DDW two
        times and collect rinses.  Quantitatively  transfer and combine
        rinses and filtrate into a  volumetric flask (50 or 100 ml
        volume) and dilute with DDW to the volume  mark.
k.      Dilute the solution from Step 3 further, if necessary.   Analyze
        metal(s) by atomic absorption spectrophotometry according to the
        instrument manufacturer's operating  instructions.  Calculate and
        report the concentration of metal(s)  in the sludge sample on a
        mg/kg dry weight basis.
                                3-10U

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                      Table 3-16
               HF-HClOij-HNOa Digestion1 **
                           Procedure
Accurately weigh a 0.5- to 1.0-g dry weight equivalent of the
homogenized sample using an analytical balance.  Transfer the
sample to a 50-ml polypropylene beaker.
Add 5 ml 48 percent hydrofluoric acid  (HF) and heat on a steam
bath at about 100C to dryness (8 to 12 hr).
Transfer residue to a 100-ml Kjeldahl flask.  Add 10 ml diges-
tion solution (5 parts cone. HNOa and 3 parts 60 percent HCTOit).
Heat on an Aminco (American Instrument Company) micro Kjeldahl
unit until the evolution of HClOij fumes.  This step should be
performed in an appropriate fume hood and a trap should be
established to catch the HCIO^ fumes.
Add 5 ml cone. HC1 and heat for 1 hr.
Cool the sample.  Dilute to approximately 30 ml with distilled
water.
Filter to remove any solid residue and dilute to 100 ml or some
other convenient volume with distilled water.  Analyze by the
method of choice.
                        3-105

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                              Table 3-17
                      HF-HNOa-HCl Digestion8'
                                   Procedure
1.      Accurately weigh 0.1- to 0.5-g dry weight equivalent  of homog-
        enized sediment using an analytical balance.  Transfer  to a
        PTFE bomb (Pan ^7^5 acid-digestion bomb or equivalent;  Pan
        Instrument Company; Moline,  Illinois).
2.      Add 6 ml W percent HF and 1 ml aqua  regia (3:1 HC1:HN03).   Seal
        the bomb and heat at 110C for 2 hr.
3.      Cool the samples and transfer to a 125-ml polypropylene wide-
        mouthed bottle containing U.8 g boric acid.
k.      The digestate can be analyzed for metals except mercury by
        transferring to a volumetric flask, adding 10 ml hydroxylammonium
        sulfate-6 percent m/v sodium chloride), and diluting  to volume.
        Analyze by method of choice.
5.      To analyze the digestate for mercury, cool the  sample bomb
        in an ice-water bath.  Carefully add  10 ml 6 percent  m/v
        potassium permanganate and let stand  30 min.
6.      Add 5 ml 5 percent potassium persulfate and allow samples to
        digest overnight at room temperature.
7-      Transfer to a volumetric flask.  Add  10 ml hydroxylammonium
        sulfate-sodium chloride solution (6 percent m/v hydroxylammonium
        sulfate-6 percent m/v sodium chloride)  and dilute to  volume.
        Analyze by method of choice.
                                3-106

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Cr, Ni, Mg, and Fe; while low temperature ashing produced low results
for As, Ni, Mg, and K.  Based on the above study, the recommended
digestion technique would be the HNOs-HCl procedure (Table 3-11).   The
HN03 procedure (Table 3-12) or the HN03-H202 procedure (Table 3-13)
would be a good second choice.
             The user is cautioned that no single digestion method may
be appropriate for all samples.  Rosengrant15  observed generally
better recoveries using l:k HC1:HN03 rather than 1:1 HClrHNOs  but did
not use the 3:1 HC1:HN03 specified by Delfino and Enderson.6   Other
factors15 that may contribute to variable results during the digestion
procedure are (l) the length of digestion period, (2) the specific
metal of interest, and (3) the type of sediment.  Longer digestion
times favor more complete metal recovery and times of 616 and 2^15
hr have been found to be convenient and useful.   Metals that have been
shown to yield variable results are Cr, Ni, Fe,  and Mn.  5  Available
data also suggest that physical factors such as  sample particle  size
or mineral composition can affect the efficiency of a digestion
procedure.15'l7
             The technical literature indicates  that the most commonly
used and reproducible digestion procedure is a variation of the  HC1-
HN03 digestion technique.6'11*"17 However, if there are problems  with
reproducibility (overall precision of +_ 10 percent or better should be
attainable6'15), metal recovery, or sample composition,  there are more
severe digestion techniques available.  These procedures involve the
use of hydrofluoric acid and/or perchloric acid.  Hydrofluoric acid
is useful to break down silicate matrices and perchloric acid is
useful for decomposing organic matter.  Variations of the HF and
HC10,, digestion procedures are presented in Tables 3-l6 and 3-17.
Unfortunately, these acids can be highly corrosive and potentially
explosive and their use is not recommended unless appropriate safety
equipment is in use in the laboratory.
Quantification procedure
             Prepare the standards in an appropriate acid medium
(depending on the digestion procedure used) or carry standards
                                3-107

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through the digestion procedure.   These solutions  should be prepared
fresh on the day of use.
             Install the appropriate hollow cathode lamp in the  instru-
ment.  Align the lamp and set the source current according  to the manu-
facturer's instructions.   Turn on the instrument and allow  both  the
instrument and the lamp to warm up.   This process  usually requires
10 to 20 min.
             Set the wavelength dial according to  Table 3-9.   The
information in Table 3-9 should only be used as a  guide in  setting up
the instrument.  Due to calibration  differences, the actual wavelength
should be based on maximum sensitivity after the instrument has  com-
pletely warmed up.  Set the slit width according to manufacturer's
instructions.
             Install the burner head indicated in  Table 3-9-
             Turn on appropriate gases, ignite flame,  and adjust the
flow of fuel and oxidant  to give maximum sensitivity for the metal
being measured.  When using a nitrous oxide flame, a T-junction  valve
or alternate switching valve should  be employed for rapidly changing
from nitrous oxide to air to prevent flashbacks when the flame is
turned on or off.
             Atomize distilled water acidified with 1.5 nil  concentrated
HNOa/ and check the aspiration rate for 1 min. If necessary, adjust
the aspiration rate to 3  to 5 ml/min.   Zero the instrument.
             Atomize a standard and  adjust the burner  alignment
(.up, down, sideways) until a maximum signal response is obtained.
             Aspirate a series of metal standards  and  record the
absorbance.  Rinse the atomizer between each standard  with  the same
acidified solution used to zero the  instrument.
             Atomize the  digested sediment samples (SID, S2,  S3) and
determine their absorbances.   Rinse  the atomizer with  dilute nitric
acid between each sample.
             When determining metal  concentrations by  atomic  absorption,
the following sequence of sample processing is recommended:
             a.  Run a set of standards.
                                3-108

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             b_.  Run five samples.
             c_.  Run a duplicate of the fifth sample.
             d_.  Run five additional samples.
             e_.  Run a duplicate of the fifth sample.
             f_.  Run a fifth sample that has been spiked.
             g_.  Run a standard.
             L.  Repeat Steps a_ through f_.
             i,.  Rerun standards.
This sequence will incorporate a quality control program into the
sample processing routine and allow the instrument operator to check
for instrument stability.
Calculations
             Prepare a standard curve by plotting the absorbance of
each standard versus concentration for each metal.  Use the standard
curve to convert sample absorbance to metal concentration.
                                3-109

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                                METALS
                                (Arsenic)

              Arsenic  is  determined using a variation of the atomic
 absorption technique.  One method consists of reduction of the arsenic
 in a sample followed  by  quantification of evolved arsine.  This offers
 the advantage of minimizing matrix interference effects during analysis
 but should be used cautiously because of the potential hazard associated
 with arsine.   The second available method for arsenic is the graphite
 furnace variation.

                      Sample Handling and Storage

              Arsenic  samples can be handled in the same manner as most
 metal samples.  Either glass or plastic containers are acceptable and
 sample integrity can  be maintained up to 6 mo with the use of nitric
 acid (Figure 3-8).

              Procedures for Water Samples (Wl, W2, S1A)

 Method 1:   Arsine Generation5'6
 Apparatus
 Arsine generator as shown in Figure 3-9.   This will include:
      a_.  Flow meter capable of measuring 1 &/min.
      b_.  Medicine dropper capable of delivering 1.5 ml and fitted
          into a size "0" rubber stopper.
      c_.  Reaction flask, which is a pear-shaped vessel with a side
          arm and a 50-ml capacity.   Both arms should have a Ik/20
          ground glass joint.
      d_.  Special gas inlet-outlet tube constructed from a micro cold
          finger condenser with the portion below the ground glass
          joint cut  off.   A Scientific Glass JM-3325 condenser or
          equivalent is suitable.
      e_.  Drying tube consisting of a 100-mm-long polyethylene tube
          filled with glass wool.
Magnetic stirrer
 References for this procedure are found oh page  3-136.
                                3-110

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               Flow Meter
        JM-3325
Medicine
Dropper in
Size "0"
Rubber
Stopper
                                      Drying Tube
                                                            (Auxi1iary Alr)
                                                              Argon
                                                            (Nebulizer Air)
                                       Hydrogen
                                         (Fuel)
                    \JM-5835
    Figure 3-9.    Arsine generator  for  arsenic  and selenium analysis
                                  3-111

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Atomic absorption spectrophotometer
Arsenic hollow cathode lamp

Reagents
Potassium iodide:  dissolve 20 g KI in 100 ml deionized distilled water.
      Prepare fresh daily.
Stannous chloride solution:  dissolve 100 g SnCl2 in 100 ml concentrated
      hydrochloric acid.
Zinc slurry:  add 50 g 200 mesh zinc metal dust to 100 ml deionized
      distilled water.
Standard diluent:  add 100 ml 18 N HaSOi* and 1*00 ml concentrated HC1
      to hOO ml deionized distilled water in a 1-H flask and dilute
      to volume with, deionized distilled water.
Perchloric acid, 70 to 72 percent.
Concentrated hydrochloric acid.
Concentrated nitric acid.
18 N_ sulfuric acid:  dilute 500 ml sulfuric acid to lwith deionized
      distilled water.
Stock arsenic solution:  dissolve 1.3209 g arsenic trioxide, AsaOa, in
      100 ml distilled water containing k g NaOH and dilute to 1 H
      with deionized distilled water.  1.00 ml = 1.00 mg As.
Intermediate arsenic solution:  pipet 1 ml stock arsenic solution into
      a 100-ml volumetric flask and dilute to volume with deionized
      distilled water containing 1.5 ml concentrated HNOs/8.
      1.00 ml = 10 yg As.
Working arsenic solution:  pipet 10 ml intermediate arsenic solution
      into a 100-ml volumetric flask and dilute to volume with
      deionized distilled water containing 1.5 ml concentrated
      HN03/2.  1.00 ml = 1 yg As.
Procedure
             Treat the sample with hydrochloric acid to determine
inorganic arsenic (a) or nitric and sulfuric acid to determine total
arsenic (/b_) :
             Inorganic Arsenic
             a..  Pipet 2 5 -ml Wl, W2, or S1A sample into a 50-ml
                 volumetric flask.  Add 20 ml concentrated hydro-
                 chloric acid and dilute to volume with deionized
                 distilled water.
                                3-112

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             Total Arsenic
             b_.  (l)  Pipet a 50-ml Wl, W2, or S1A sample into a 150-ml
                      "beaker.  Add 10 ml concentrated nitric acid and
                      12 ml 18 N sulfuric acid.
                 (2)  Evaporate the sample to the evolution of SOa fumes
                      (approximately 20 ml).  To avoid the loss of
                      arsenic, add small amounts of nitric acid whenever
                      the red-brown NOa fumes disappear.
                 (.3)  Cool slightly, add 25 ml deionized  distilled water
                      and 1 ml perchloric acid, and evaporate to SOa
                      fumes.  Cool, add kd ml concentrated hydrochloric
                      acid, and dilute to a volume of 100 ml with
                      deionized distilled water.
             Prepare standard arsenic solutions by diluting 0, 0.5,
1.0, 1.5S and 2.0 ml of working arsenic solution to 100 ml with the
standard acid diluent.  These solutions contain 0, 5, 10, 15s and 20  Vg
As/&.
             Pipet 25 ml of the sample [either (a_) or (b_)] or standard
arsenic solution into the reaction vessel.  Add 1.0 ml KI solution and
0.5 ml SnCla solution.  Allow 10 min for the arsenic to be reduced to
the lowest oxidation state.
             Attach the reaction vessel to the special gas inlet-outlet
glassware.  Fill the medicine dropper with 1.50 ml homogenized zinc
slurry and insert the medicine dropper in the side neck of the reaction
vessel.  With the arsine generator attached to the atomic absorption
spectrophotometer using an argon-hydrogen flame, add the  zinc slurry
to the sample.   The arsine peak should occur almost immediately.   When
the recorder pen returns part way to the baseline, remove the reaction
vessel.
Calculations
             Prepare a standard curve by plotting peak height versus
arsenic concentration in the standards.  Determine the arsenic con-
centration by comparing sample peak height with the standard curve.
             It is necessary to multiply the determined sample
arsenic concentrations by a factor of 2 to correct for the fact that
samples were diluted by 50 percent with acid and the standards were
not.
                                3-113

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             Report results with Wl samples as total arsenic and
results with W2 and S1A samples as soluble arsenic.
             If it is necessary or desirable to determine organic
arsenic, this can "be calculated as total arsenic (digestion b_)  minus
inorganic arsenic (digestion a_) .
Method 2:  Graphite Furnace

Apparatus

Atomic absorption spectrophotometer equipped with a graphite furnace
      or carbon rod atomizer and background corrector

Recorder

Automatic sampler or Eppendorf pipettes

Hot plate or hot water bath

Reagents

Stock solution:  dissolve 1.320 g of arsenic trioxide, As20a (analy-
      tical reagent grade), in 100 ml of deionized distilled water
      containing it g NaOH.  Acidify the solution with 20 ml cone.
      HN03 and dilute to 1 i.  1 ml = 1 mg As.

Working arsenic solution:  prepare dilutions of the stock solution to
      be used as calibration standards at the time of analysis.  With-
      draw appropriate aliquots of the stock solution and add 1  ml of
      concentrated HN03, 2 ml of 30 percent H2C>2, and 2 ml of the
      5 percent nickel nitrate solution.  Dilute to 100 ml with
      deionized distilled water.

Nickel nitrate solution, 5 percent:  dissolve 2^.780 g of ACS reagent
      grade Wi(NOa)2  6H20 in deionized distilled water and make  up
      to 100 ml.

Nickel nitrate solution, 1 percent:  dilute 20 ml of the 5 percent
      nickel nitrate to 100 ml with deionized distilled water.

Procedure

             Transfer a 100-ml Wl, W2, or S1A sample to a 250-ml
Griffin beaker.  Add 2 ml 30 percent HaOa and sufficient concentrated
HN03 to produce a 1 percent (v/v) acid concentration (approximately
1 ml).  Heat for 1 hr at 95C or until the volume is slightly less
than 50 ml.
             Cool and dilute the sample to 50 ml with deionized
distilled water.
             Pipet 5 ml of digested sample to a 10-ml volumetric

-------
flask.  Add 1 ml of 1 percent nickel nitrate solution and  dilute to
10 ml with deionized distilled vater.
             Inject 20-yl aliquots of standard and digested  samples.
Record the sample absorbance.  The method of standard additions
should be used to quantitate the sample unless it  can be shown that
the sample matrix does not affect the results.10
Calculations
             Prepare a standard curve by plotting  absorbance of the
arsenic standards versus arsenic concentration.  Determine the
arsenic concentration of the samples by comparing  the observed
absorbance with the standard curve.
                               3-115

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             Procedure for Sediment Samples (SID, S2,  S3)

Method 1:  Arsine Generation11
Apparatus
Atomic absorption spectrophotometer
Arsine generator as shown in Figure 3-9 and described  in the procedure
      for the analysis of arsenic in water samples.  Use a 5-ml
      medicine dropper rather than a 1.5-ml medicine dropper
Reagents
Concentrated hydrochloric acid.
Potassium iodide:  dissolve 15 g in 100 ml distilled deionized water.
      Prepare daily.
Stannous chloride:  dissolve ko g SnCla in 100 ml concentrated hydro-
      chloric acid.
Zinc slurry:  add 50 g 200 mesh zinc metal dust to 100 ml  deionized
      distilled water.
Stock arsenic solution:  dissolve 1.320 g arsenic trioxide,  As20.3,  in
      10 ml deionized distilled water containing U g NaOH.   Dilute  it
      to 1  with deionized distilled water.   1.00 ml  = 1.00 mg As.
Intermediate arsenic solution:  pipet 1 ml stock arsenic solution  into
      a 100-ml volumetric flask and dilute to volume with deionized
      distilled water containing 1.5 ml concentrated HN03/.  1.00  ml  =
      10 yg As.  Prepare arsenic standards daily in the appropriate
      range.
Procedure
             Weigh out 0.5 g dry weight equivalent of  the  sediment
sample (SID, S2, S3).  Transfer to a lOO^nl beaker and add 2.5 g
potassium pyrosulfate.  If larger sediment samples are used, propor-
tionately increase the amount of potassium pyrosulfate used.
             Fuse the sample at 320C for 15 min in a  furnace.  Cool
the sample.
             Dissolve the residue in 25 ml deionized distilled water
and 5 ml concentrated hydrochloric acid.  Heat the sample  in a water
bath, if necessary, to dissolve the solids.
             Transfer the sample to a 100-ml volumetric flask and
dilute to volume with deionized distilled water.
             Pipette 25 ml of sample or standard to the reaction
vessel in Figure 3-8.  Add 5 ml concentrated hydrochloric  acid and

                                3-116

-------
mix thoroughly.  Add 2 ml potassium iodide solution and  mix thoroughly.
Add 0.5 ml stannous chloride and allow 30 min for  the  reduction  of
arsenic.
             Attach the reaction vessel to the special glass inlet-
outlet tube of the arsenic generator attached to the atomic absorption
spectrophotometer.
             While stirring the zinc slurry,  fill  the  medicine dropper
with 5 ml of zinc slurry.  Insert the medicine dropper into the  side
neck of the reaction vessel.  When the instrument  is warmed up and
stable, add the zinc slurry to the sample. The arsenic  peak should
be detected almost immediately.  A hydrogen-argon  flame  should be
used.   When the recorder response has approached baseline,  remove the
reaction vessel and replace with the next sample.
Calculations
             Prepare a standard curve based on the absorbance and
concentration of the arsenic standards.  Determine the arsenic
concentration in the sediment digests by comparing the sample
absorbance with the standard curve.   Calculate the arsenic  concentra-
tion of the sediment sample as follows:
                   /.   /  ,    . ,.N    (x) (0.1 &)  (1000)
              As yg/kg (wet weight)  = 		
                                               D
              As pg/kg (dry weight)  - '*' ((s} \\  ^
where
         x = the arsenic concentration in the sediment digest, yg/C
     0.1 H = the final sample volume
         g = the weight of wet sediment digested,  g
       % S = the percent solids in the sediments as a  decimal fraction
                               3-117

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                                METALS
                               (.Mercury)

             Mercury is also determined using a variation of the atomic
absorption technique.  However, due to the fact that a different diges-
tion solution is used and the fact that mercury is quantified using a
cold vapor technique rather than conventional flame atomic absorption
spectrophotometry, mercury is discussed independent of the other metals.

                      Sample Handling and Storage

             Samples for mercury analysis should preferably be stored
in glass containers because it has been shovn that polyethylene con-
tainers are permeable to mercury vapors.     The recommended method
of sample preservation10 is to add ni'tric acid to a pH of 2 or less.
The recommended holding time for samples preserved in this manner is
38 days for samples in glass and 13 days for samples in plastic
(Figure 3-10).  However, it has been reported19'20 that the addition
of potassium dichromate and nitric acid is a better preservative than
nitric acid alone.  In either case, hydrochloric acid should not be
used as a preservative since mercury may be lost as the volatile
mercury chloride.
             The use of moist sediment samples is recommended.   If
samples are dried or frozen, care should be taken not to use
excessively high temperatures during drying or thawing as this may
result in the loss of volatile mercury compounds (Figure 3-10).

               Procedure for Water Samples (Wl, W2, S1A)
Method 1:  Cold Vapor Technique5'10
Apparatus
Atomic absorption spectrophotometer  equipped with a glass  cell  as
      schematically indicated in Figure 3-11. Alternately,
_                                                     _
  References for this procedure are  found on page 3-136.
                                3-118

-------

C

WATER SAMPLE DREDGE SAMPLE
x 4

ACIDIFY FILTER N TR"ENT STORE WET
1 1 	
STORE ACIDIFY
1 1
i '
1' ' +
DIGEST STORE 1 | ELUTRIATE FRACTIONATE B'S1C?Y
11 11 	
ANALYZE ANALYZE ANALYZE ANALYZE
(W1) (W2) (S1A) (S1B)
LO
VQ1 SAMPLE DESIGNATION
PURPOSE
CONTAINER
SAMPLE TREATMENT
PRESERVATIVE
ORE SAMPLE
1

CORE SECTION
i
+
DRY
1
STORE


DIGEST DIGEST
1 1
ANALYZE ANALYZE
(SID) (52)

W1 W2 W3 S1A SIB SIC SID S2


FREEZE
1
STORE
I
DIGEST
1
ANALYZE
(S3)

S3


Total Water Soluble Used in Mobile Chemical Bloavall- Total Total Total
Cone. Water Elutriate Cone. Distribution ability Sediment Sediment Sediment
Cone. Cone. Cone. Cone.
GGG GGGGGG
None Filter None None None None None (<60C) Freeze
HN03 HN03 None I(C 
-------
Figure 3-11.    Schematic cold vapor apparatus for mercury
^
1
1
,F
H
                                      B
                                    LEGEND;
                                    A -  Reaction  Flask
                                    B-  Drying  tube,  filled with MgClOi,
                                    C-  Rotameter, 2  liters of air  per
                                         minute
                                    D-  Absorption cell with quartz
                                         windows
                                    E-  Compressed air, 2  liters of air
                                         per minute
                                    F -  Glass tube with fritted end
                                    G-  Hollow  cathode mercury lamp
                                    H -  AA detector
                                    J -  Vent to hood
                                    K-  Recorder, any compatible model
                              3-120

-------
      commercially available cold vapor technique instruments
      specifically designed for mercury analysis may be substituted
      for the atomic absorption spectrophotometer.   The flow cell
      should be approximately 2.5 cm in diameter, as long as the
      instrument will permit, and have quartz windows on each  end.
      Support the cell in the light path of the instrument to  give
      maximum transmittance

Mercury hollow cathode lamp

Air pump capable of delivering 1 to 2  air/min

Flow meter capable of measuring 1 to 2 5,/min

Aeration tubing:  a straight glass frit having a coarse porosity.
      Tygon tubing is used for passage of the mercury vapor from the
      sample bottle to the absorption cell and through the apparatus

Drying tube:  150 mm x 18 mm diameter containing 20 g magnesium
      perchlorate, MgClOit
Reagents

Sulfuric acid, cone:  reagent grade.

Sulfuric acid, 0.5 N_:  dilute 1^.0 ml of cone, sulfuric acid to 1.0 &.
Nitric acid, cone.:  reagent grade of low mercury content.
NOTE:  If a high reagent blank is obtained, it may be necessary to
       distill the nitric acid.
Sodium chloride-hydroxylamine sulfate solution:  dissolve 12 g of
      sodium chloride and 12 g of hydroxylamine sulfate in distilled
      water and dilute to 100 ml.  (Hydroxylamine hydrochloride may
      be used in place of hydroxylamine sulfate.)
Potassium permanganate:  5 percent solution, w/v.  Dissolve 5  g of
      potassium permanganate in 100 ml of distilled water.

Potassium persulfate:  5 percent solution, w/v.  Dissolve 5 g  of
      potassium persulfate in 100 ml of distilled water.

Stock mercury solution:  dissolve 0.135^ g of mercuric chloride in
      75 ml of distilled water.  Add 10 ml of cone, nitric acid and
      adjust the volume to 100.0 ml.  1 ml = 1 mg Hg.

Stannous chloride:  dissolve 100 g SnCl2 in deionized distilled water
      containing 12.5 ml concentrated HC1 and dilute to 1  with
      deionized distilled water.  Stir continuously during use if a
      suspension forms.

NOTE:  A stannous sulfate solution may be prepared and used in place
       of the stannous chloride solution.

Working mercury solution:  make successive dilutions of the stock
       mercury solution to obtain a working standard containing 0.1 g
       per ml.  This working standard and the dilutions of the stock
       mercury solution should be prepared fresh daily.  Acidity of
                                3-121

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      the working standard should be maintained at 0.15 percent nitric
      acid.  This acid should be added to the flask as needed before
      the addition of the aliquot from the stock mercury solution.
Procedure
             Transfer a 100-ml sample of each standard mercury solution
and each Wl, W2, and S1A sample to a 300-ml BOD bottle.  Add 5 ml con-
centrated HaSOit and 2.5 ml concentrated HNOs to each flask.   Add 15 ml
potassium permanganate solution to each flask and let stand a minimum
of 15 min.  Add 8 ml potassium persulfate solution to each flask and
heat in a water bath at 95C for 2 hr.  Following digestion, cool the
samples to room temperature.
             While the samples are digesting, set up the instrument
according to the manufacturer's instructions.  This would include:
             a_.  Install and align the hollow cathode lamp.
             b_.  Set the wav.elength at 253.7 nm.
             c_.  Set the slit width and lamp current at recommended
                 values.
             d..  Allow the instrument to warm up.
             e_.  Install the mercury absorption cell.
             .  Adjust the air flow to 2 &/min.
             Add 6 ml sodium chloride-hydroxylamine sulfate to the
cooled sample digest.  Allow at least 5 min to reduce any excess
permanganate.  From this point on, each standard and sample must be
treated individually to completion.
             Add 5 ml stannous chloride and immediately attach the BOD
bottle to the aeration apparatus.  The maximum absorbance should occur
within a few seconds.
             As soon as the recorder returns approximately to the
baseline, remove the stopper holding the frit from the reaction flask'
and replace the sample bottle with a bottle containing deionized
water.  Flush the system for a few seconds and run the next sample/
standard.
             Because of the toxic nature of mercury vapor, care must
be taken to avoid its inhalation.  Therefore, the mercury vapor should
be vented to an exhaust hood or through an absorbing media such as
                                3-122

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(a) O.i M KMnOij and 10 percent H2S04 or (b)  0.25  percent  iodine  in a
3 percent KI solution.
Calculations
             Prepare a standard curve by plotting absorbance  of  the
mercury standards versus mercury concentration.   Determine  the mercury
concentration of the samples by comparing the sample  absorbance  with
the standard curve.
             Report the Wl sample results as total mercury  and the W2
and S1A results as soluble mercury.
                                3-123

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               Procedure for Sediment Samples  (SID, S2, S3)


Method 1:  Cold Vapor Technique11'21'22

Apparatus

Atomic absorption spectrophotometer equipped with a glass cell and an
      aeration apparatus as shown in Figure 3-11.  Alternately,
      commercially available cold vapor  instruments designed specifically
      for mercury may be used

Mercury hollow cathode lamp

Air pump capable of delivering 2  air/min

Flow meter capable of measuring 2 5,/min

Aeration tubing:   a straight glass frit  having a coarse porosity.
      Tygon tubing is used for passage of the mercury vapor from the
      sample bottle to the absorption cell and through the apparatus

Drying tube:  150 mm x 18 mm diameter containing 20 g magnesium
      perchlorate, MgClOi,

Reagents

Sulfuric acid, cone.:  reagent grade.

Nitric acid, cone.:  reagent grade of low mercury content.

NOTE:  If a high reagent blank is obtained, it may be necessary to
       distill the nitric acid.
Sodium chloride-hydroxylamine sulfate solution:  dissolve 12 g of
      sodium chloride and 12 g of hydroxylamine sulfate in distilled
      water and dilute to 100 ml.  (Hydroxylamine hydrochloride may
      be used in place of hydroxylamine  sulfate.)

Potassium permanganate:  5 percent solution, w/v.  Dissolve 5 g of
      potassium permanganate in 100 ml of distilled water.

Potassium persulfate:  5 percent solution, w/v.  Dissolve 5 g of
      potassium persulfate in 100 ml of  distilled water.
Stock mercury solution:  dissolve 0.135^ g of mercuric chloride in
      75 ml of distilled water.  Add 10  ml of cone, nitric acid and
      adjust the volume to 100.0 ml.  1  ml = 1 mg Hg.

Prepare working mercury solutions by appropriate dilution of the stock
      mercury solution on the day of use.

Stannous chloride:  dissolve 100 g Snda in deionized distilled water
      containing 12.5 ml concentrated HC1 and dilute to 1 H with
      deionized distilled water.  Stir continuously during use if a
      suspension forms.

NOTE:  A stannous sulfate solution may be prepared and used in place
       of the stannous chloride solution.10
                                3-121;

-------
Procedure
             Set up the instrument according to the manufacturer's
instructions.  This should include:
             a_.  Installing and aligning the hollow cathode lamp.
             b_.  Setting the lamp wavelength at 253.7 nm.
             c_.  Setting the slit width and lamp current at recommended
                 values.
             d_.  Allowing the instrument to warm up for 10 to 20 min.
             e_.  Installing the mercury absorption cell.
             f_.  Adjusting the air flow to 2 i/min.
             Homogenize the sediment sample and weigh out a 0.5 to 2 g
dry weight equivalent of the moist sample.  Transfer the sample to a
300-ml BOD bottle and rinse the sediment to the bottom of the flask
with distilled deionized water.
NOTE:  Dried or frozen sediment samples may be used but moist samples
       are recommended since their use avoids the possible loss of
       volatile mercury compounds during the drying or thawing cycles.
             Cool the sample in an ice bath and add 5 ml concentrated
sulfuric acid and 5 ml concentrated nitric acid.  The use of the ice
bath is intended to counteract the heating of the sample that can
result from the addition of the acid and the potential volatilization
of mercury that may result.
             Add 15 'ml 5 percent potassium permanganate.  If the pink
permanganate color does not persist for 15 min, add additional per-
manganate.  Digest the unstoppered sample for 2 hr in a 60C water bath.
             Cool the sample and add 5 ml potassium persulfate solution.
Stopper the samples and allow to stand overnight.  Add sufficient
hydroxylamine sulfate-sodium chloride solution until the brown
hydrated manganese oxides and excess potassium permanganate color  are
dissipated.  Add approximately 100 ml distilled water.
             Add 10 ml stannous chloride to the sample and immediately
attach the sample to the aeration apparatus.  The maximum absorbance
should occur within a few seconds.
             As soon as the recorder returns approximately to the
baseline, remove the stopper holding the frit from the reaction flask
                                 3-125

-------
and replace the sample bottle with a bottle containing deionized water.
Flush the system for a few seconds and run the next standard/sample.
             Because of the toxic nature of mercury vapors,  the sample
should be vented to an exhaust hood or through an absorbing  media such
as (a) 0.1 M KMn(X and 10 percent HaSO"* or (b) 0.25 percent  iodine in
3 percent KI.
Calculations
             Prepare a standard curve based on the absorbance of the
mercury standards and the amount of mercury in the prepared  standards.
Compare the sample absorbance to the standard curve to determine the
amount of mercury in the sample digest.  Calculate the mercury con-
centration in the sediment as follows:
                        /,   /  ,    .  ,.\    1000 x
                    Hg yg/kg (wet weight)  = -
                                               &
                    Hg yg/kg (dry weight)  =       X
where
       x = weight of mercury in the sample,  yg
       g = wet weight of sediment used,  g
     % S = percent solids in the sediment sample as  a decimal  fraction
                                3-126

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                                METALS
                              (Selenium)

                      Sample Handling and Storage

             Selenium samples can be preserved with nitric acid as indi-
cated for most other metals (Table 3-8).  This method is considered
acceptable for vater samples up to periods of 6 mo.  Glass or plastic
containers should be acceptable with sediment samples since both are
acceptable with water samples.  However, the time limits for sediment
storage are not known.

               Procedure for Water Samples (Wl, W2, S1A)5

Method 1:  Hydride Generation
Apparatus
Hydride generator as diagramed in Figure 3-9  This will include:
      a_.  Flow meter, capable of measuring 1 &/min
      b_.  Medicine dropper, capable of delivering 1.5 ml, fitted into
          a size "0" rubber stopper
      c_.  Reaction flask, which is a pear-shaped vessel with side arm
          and a 50-ml capacity.  Both arms should have a 14/20 ground
          glass joint
      cl.  Special glass inlet-outlet tube constructed from a micro
          cold finger condenser with the portion below the ground
          glass joint cut off.  A Scientific Glass JM-3325 or equi-
          valent is suitable
      e_.  Drying tube consisting of a 100-mm-long polyethylene tube
          filled with glass wool
Magnetic stirrer
Atomic absorption spectrophotometer
Hollow cathode lamp
Reagents
Stannous chloride:  dissolve 100 g SnCla in 100 ml concentrated
      hydrochloric acid.
Zinc slurry:  add 50 g 200 mesh zinc metal dust to 100 ml deionized
	distilled water.	
 References for this procedure are found on page 3-136.
                                3-127

-------
Standard diluent:  add 100 ml 18 N HaSO"*  and ^00 ml concentrated HC1 to
      kOQ ml deionized distilled water in a 1-& volumetric flask and
      dilute to volume with deionized distilled water.

Stock selenium solution:  dissolve 1.000 g selenium in 5 ml concentrated
      HNOs.   Warm until the reaction is complete and cautiously evaporate
      just to dryness.  Dilute to 1 & with deionized distilled water.
      1.00 ml = 1.00 mg Se.  (Alternately dissolve 0.3^53 g selenous
      acid (assay ^.6 percent HaSeOs) in deionized distilled water and
      dilute to 200 ml.  1.00 ml = 1.00 mg Se.

Intermediate selenium solution:  pipet 1 ml stock selenium solution into
      a 100-ml volumetric flask and dilute to volume with deionized
      distilled water containing 1.5 ml concentrated HNOaA.   1.00 ml =
      10 Ug Se.

Working selenium solution:  pipet 10 ml intermediate selenium solution
      into a 100-ml volumetric flask and dilute to volume with
      deionized distilled water containing 1.5 ml concentrated HN03/&.
      1.00 ml = 1.00 yg Se.  Prepare fresh on the day of use.

Perchloric acid, TO to 72 percent.

Concentrated nitric acid.

18 N_ sulfuric acid:  dilute 500 ml concentrated sulfuric acid to 1 &
      with deionized distilled water.

Procedure

             Treat the sample with hydrochloric acid to determine
inorganic selenium (a) or nitric acid and sulfuric acid to determine

total selenium (b).  Organic selenium can be calculated as the dif-

ference between these two treatments.

             Inorganic Selenium
             a.  Pipet 25-ml Wl, W2, or S1A sample into a 50-ml
                 volumetric flask.   Add 20 ml concentrated hydrochloric
                 acid and dilute to volume with deionized distilled
                 water.
             Total Selenium

             b_.  (.1)  Pipet a 50-ml Wl, W2, or S1A sample into a 150-ml
                      beaker.  Add 10 ml concentrated nitric  acid and
                      12 ml 18 1J sulfuric acid.
                 (2)  Evaporate the sample to the evolution of SO3
                      fumes (approximately 20 ml).  To  avoid the loss
                      of selenium,  add small amounts of nitric acid
                      whenever the red-brown NOa fumes  disappear.

                 (3)  Cool slightly.  Add 25 ml deionized distilled
                      water, 1 ml perchloric acid, and  evaporate to
                          fumes.  Cool, add ho ml concentrated


                                3-128

-------
                      hydrochloric acid, and dilute to a volume of
                      100 ml with deionized distilled water.
             Prepare standard selenium solution by diluting 0, 0.5, 1.0,
1.5, and 2.0 ml working selenium solution to 100 ml with the standard
acid diluent.  These solutions contain 0, 5, 10, 15 and 20 yg Se/,
respectively.
             Pipet 25 ml of the sample [either (a) or (b)]  or standard
selenium solution into the reaction vessel.   Add 0.5 ml stannous
chloride solution and allow 10 min for the selenium to be reduced.
             Attach the reaction vessel to the special gas inlet-
outlet glassware.  Fill the medicine dropper with 1.5 ml homogenized
zinc slurry and insert the medicine dropper  into the side neck of the
reaction vessel.  With the hydride generator attached to the atomic
absorption spectrophotometer and the instrument producing a stable
response with an argon-hydrogen flame, add the zinc slurry to the
sample.  The selenium hydride peak should occur almost immediately.
When the recorder pen returns part way to the established baseline,
remove the reaction vessel.
             Continue processing samples and standards in a similar
manner.
             The following general procedure should be used when
processing samples:
             a_.  Run a set of standards.
             b_.  Run five samples.
             c_.  Run a duplicate of the fifth sample.
             <1.  Run five samples.
             e_.  Run a duplicate and a spike of the fifth sample.
             f_.  Run a standard.
             g_.  Repeat the cycle.
In this way, a quality control program can be incorporated into the
sample processing routine and the instrument stability can be checked.
Calculations
             Prepare a standard curve by plotting standard absorbance
versus selenium concentration in the standards.  Determine the
selenium concentration by comparing the sample absorbance with the
                                 3-129

-------
standard curve.  Be sure to multiply the determined selenium concentra-
tions "by 2 as the samples vere diluted 1:1 with acid and the standards
were not.
             Report results for Wl samples as total selenium and results
for W2 and S1A samples as soluble selenium.
             If it is necessary or desirable to determine organic
selenium, this can be calculated as total selenium (digestion b)
minus inorganic selenium (digestion a).
                                 3-130

-------
            Procedures for Sediment Samples (SID, S2, S3)

                                                f\ n
Method 1:  Digestion/Flameless Atomic Absorption
Apparatus
Atomic absorption spectrophotometer equipped with a deuterium background
      corrector
Selenium electrodeless discharge lamp:  operate at 9 watts, a slit width
      of 0.7 and 196.0 nm
Automatic sampler
Eppendorf microliter pipets may be used where an automatic sampler is
      not available
Graphite furnace:  dry samples at 125C, char samples at 1500C for 30
      sec, and atomize samples for 10 sec at 2700C
Hot plate
Reagents
Concentrated nitric acid, redistilled.
Hydrogen peroxide, 30 percent.
Stock selenium solution:  dissolve 0.3^53 g selenous acid (.assay
      9^.6 percent HaSeOs) in deionized-distilled water and dilute
      to 200 ml.  1.00 ml = 1.00 mg Se.
Intermediate selenium solution:  pipet 1 ml stock selenium solution
      into a 100-ml volumetric flask and dilute to volume with
      deionized distilled water containing 1.5 ml concentrated
      nitric acid/A.  1.00 ml = 10 Pg Se.
Prepare selenium standards in the appropriate range on the day of use.
      Add the selenium to a 100-ml volumetric flask containing 1 ml
      concentrated nitric acid and 2 ml 30 percent  hydrogen peroxide.
      Dilute to volume with deionized distilled water.
1 percent nickel nitrate:  dissolve ^.956 g Ni(N03)2  6 HaO in 100 ml
      deionized distilled water.
Nickel nitrate, 5 percent:  dissolve 2^.780 g Ni(NOs)2  6 HaO in 100 ml
      deionized distilled water.
Procedure
             Dry the sample to be analyzed at 60C.  Temperatures
above this value are not recommended due to the possibility of selenium
loss as a result of volatilization.  Weigh a 0.5-g  sample of the dried
material (SID, S2, S3) and transfer to a 250-ml Griffin beaker.
             Add 5 ml concentrated HNOs to the sample and cover the
beaker with a watch glass.  Reflux the sample to near dryness at 95C.

                                 3-131

-------
After the sample has cooled, add a second 5-ml portion of concentrated
nitric acid and repeat the digestion.   Cool the sample.
             Add 3 ml concentrated nitric acid and 10 ml 30 percent
hydrogen peroxide.  Place the beaker on a hot plate and warm gently
until a reaction commences.  Immediately remove the "beaker from the  hot
plate until the vigorous effervescence has subsided.   Return the covered
beaker to the hot plate and reflux the sample at 95C for an additional
15 min.
             Cool the sample and dilute to 50 ml with deionized
distilled water.
             Pipet 5 ml of digested sample or standard into a 10-ml
volumetric flask.  Add 2 ml 5 percent nickel nitrate solution and
dilute to volume with deionized distilled water.   Allow any particulate
matter to settle before withdrawing an aliquot for analysis.
NOTE:  Selenium standards and samples to be analyzed by the graphite
       furnace method are mixed with a nickel nitrate solution to
       enhance sensitivity.  This step will double the sensitivity
       but care must be taken to treat samples and standards in a similar
       fashion to avoid introducing a differential matrix effect.
             Because of possible matrix effects,  the method of standard
additions should be used.  Additional 5-ml aliquots of the digested
sample should be pipeted into 10-ml volumetric flasks and spiked with
known amounts of selenium standard.  Add 2 ml 5 percent nickel nitrate
and dilute to volume with deionized distilled water.
             Samples sizes of 5 to 10 yl should be injected into the
graphite furnace and dried for 20 sec, charred for 30 sec, and
atomized for 10 sec.  The small sample size is recommended to minimize
possible interference in the sediment digests.  If larger sample sizes
are used, the drying time will have to be increased.
             Record the data for standards, samples, and spiked samples.
Calculations
             With the standard addition approach, plot the absorbance
of the sample and the absorbance of the spiked samples versus the
amount of added selenium.  Extrapolate the data to determine the amount
of selenium in the sample digest.
                                3-132

-------
             When using a series of standard selenium concentrations,
plot standard absorbance versus selenium concentration.   Determine  the
selenium concentration in the sample digest by comparing sample absor-
bance with the standard curve.
                 Se Ug/kg (vet  weight)  = (x) (L)  100
                                               o
                 Se ug/kg (dry weight)  =
where
       x = selenium concentration in sample digest,
       L = final volume of sample digest,  (0.05 &  as written)
       g = wet weight of sediment digested, g
     % S = percent solids in sediment sample as a decimal fraction

Method 2:  Hydride Generation
Apparatus
Atomic absorption spectrophotometer
Hydride generator as shown in Figure 3-9 and described in the procedure
      for the analysis of selenium in water samples.   Use a 5-ml
      medicine dropper rather than a 1.5-ml medicine  dropper
Reagents
Concentrated hydrochloric acid.
Stannous chloride:  dissolve ho g SnCla in 100 ml concentrated hydro-
      chloric acid.
Zinc slurry:  add 50 g 200 mesh zinc metal dust to 100 ml deionized-
      distilled water.
Stock selenium solution:  dissolve 1.000 g selenium  in 5 ml concentrated
      HNOs.  Warm until the reaction is complete  and  cautiously evapo-
      rate just to dryness.  Dilute to  1 Si with deionized distilled
      water.  1.00 ml = 1.00 mg Se.  (Alternately, a  stock solution can
      be prepared by dissolving 0.3^53  g selenous acid (assay 9^-6  per-
      cent ^SeOa) in deionized distilled water and  diluting to 200 ml.
      1.00 ml = 1.00 mg Se.
Intermediate selenium solution:   pipet  1 ml stock selenium solution into
      a 100-ml volumetric flask and dilute to volume  with deionized
      distilled water containing 1.5 ml concentrated  HN03/.  1.00  ml  =
      10 yg Se.
Prepare selenium standards in the appropriate range  on the day of use.
                                 3-133

-------
Procedure
             Weigh a 0.5- to 1.0-g dry weight equivalent  of sediment
sample (SID, S2, S3) and transfer to a 150-ml beaker.   Add 10 ml con-
centrated nitric acid and 12 ml 18 N_ sulfuric acid.
             Cover the sample with a watch glass and heat on a hot plate
until SOs fumes are evolved.  To avoid the loss of selenium, replenish
the nitric acid whenever the red-brown N02 fumes disappear.
             Allow the sample to cool.  Add 25 ml deionized distilled
water and 1 ml perchloric acid and reheat the sample at 95C until SOa
fumes appear.  Cool the sample and add ko ml concentrated hydrochloric
acid.  Transfer the sample to a 100-ml volumetric flask and dilute to
volume with deionized distilled water.
             Prepare a series of standard selenium solutions.  Pipet
the appropriate amount of working selenium solution into a series of
100-ml volumetric flasks and dilute to volume with standard acid
diluent.
             Pipet 25 ml of digested sample or standard into the
reaction vessel.  Add 0.5 ml stannous chloride solution and allow
10 to 15 min for the selenium to be reduced.
             Attach the reaction vessel to the special gas inlet-outlet
glassware.  Fill the medicine dropper with 1.5 ml homogenized zinc
slurry and insert the medicine dropper into the side neck of the
reaction vessel.  With the hydride generator attached to the atomic
absorption spectrophotometer and the instrument producing a stable
response with an argon-hydrogen flame, add the zinc  slurry to the
sample.  The selenium hydride peak should occur almost immediately.
When the recorder pen returns part way to the established baseline,
remove the reaction vessel.  Continue processing standards and samples
in a similar manner.
Calculations
             Prepare a standard curve by plotting standard absorbance
versus selenium concentration in the standards.  Determine the selenium
concentration in the sample digests by comparing the sample absorbance
with the standard curve.
                                3-13U

-------
             Calculate the selenium concentration in  the  sediment  samples
as follows:
              e    /i   I   4.     ^\   (x)  (0.1 &)  (1000)
              Se yg/kg (wet weight) = --J	--	-
                                               g
              a    h   i A       ^\   (x)  (o.i a)  (looo)
              Se yg/kg (dry weight) =      (g) (% S)	
where
       x = selenium concentration in the sediment digest, yg/&
     0.1 = volume of sediment digest, H
       g = wet weight of sediment .digested,  g
     % S = percent solids in the sediment  sample as a decimal fraction
                                 3-135

-------
                               References
 1.  Stumm, W., and Morgan, J. J.  Aquatic Chemistry.   Wiley-Interscience;
     New York, New York.  538 p. (1978).

 2.  Williams, S. L., Aulenbach, D. B., and Clesceri,  N.  L.   "Sources
     and Distribution of Trace Metals in Aquatic Environments."  In:
     Aqueous Environmental Chemistry of Metals.   A.  J. Rubin (Ed.).
     Ann Arbor Science Publishers;  Ann Arbor, Michigan.  390 p. (197M-
     pp. 77-127.
 3.  Helz, G. R.  "Trace Element Inventory for the Northern  Chesapeake
     Bay with Emphasis on the Influence of Man."  Geochem et Cosmochem
     Acta 1*0:573-580 (1976).
 U.  Keeney, W. L., Breck, W. G., Vanloon, G. W., and  Page,  J.  A.
     "The Determination of Trace Metals in Cladophora  Glomerata -.-
     C_. Glomerata as a Potential Biological Monitor."   Water Research 10:
     981-981* (1976).
 5.  American Publich Health Association.  Standard Methods  for the
     Examination of Water and Wastewater Including Bottom Sediments
     and Sludges, l^th Edition.APHA; New York, New York.1193 p.
     (1976)7
 6.  Delfino, J. J., and Enderson,  R. E.  "Comparative Study Outlines
     Methods of Analysis of Total Metal in Sludge."  Water and  Sewage
     Works 125(RN):R32-3^, i*7-1i8 (1978).
 7.  Varian Techtron.  "Water Analysis by Atomic Absorption  Spectro-
     scopy."  Varian Techtron; Palo Alto, California.   78 p. (1976).

 8.  Environment Canada.  "Analytical Methods Manual."  Inland  Waters
     Directorate, Water Quality Branch; Ottawa,  Ontario,  Canada (197*0 .

 9.  Environmental Protection Agency.  "Water Programs. Guidelines
     Establishing Test Procedures for the Analysis of  Pollutants."
     Federal Register Ul(232):52780-52786 (Wednesday,  1 December 1976).
10.  Environmental Protection Agency.  "Methods for  Chemical Analysis
     of Water and Wastes."  Environmental Monitoring and Support
     Laboratory, EPA; Cincinnati, Ohio (1979).
11.  Walton, A.  "Ocean Dumping Report 1. Methods for  Sampling  and
     Analysis of Marine Sediments and Dredged Materials." Department
     of Fisheries and Environment;  Ottawa, Ontario,  Canada.   7^ p. (1978).
12.  Environmental Effects Laboratory.  "Ecological  Evaluation  of
     Proposed Discharge of Dredged or Fill Material  into Navigable
     Waters. Interim Guidance for Implementation of  Section  UoU(b)(l) of
     Public Law 92-500 (Federal Water Pollution Control Act  Amendments
     of 1972)."  Miscellaneous Paper D-76-17.  U. S. Army Engineer
     Waterways Experiment Station,  CE; Vicksburg, Mississippi.   83 p.
     (1976).
                                  3-136

-------
13.  Delfino, J. J., Bortleson,  G.  C.,  and Lee,  G.  F.   "Distribution of
     Mn, Fe, P, Mg, K, Na, and Ca in the Surface Sediments of Lake
     Mendota, Wisconsin."  Environmental Science and Technology 3:ll89-
     1192 (1969).
lit.  Agemien, H. , Aspila, K.  I., and Chau, A.  S.  Y.   "A Comparison of the
     Extraction of Mercury from Sediments by Using  Hydrochloric-Nitric
     Acid, Sulfuric-Nitric and Hydrofluoric Acid-Aqua Regia Mixture."
     Analysis 100:253-258 (1975).
15.  Rosengrant, L. E.  "A Method Study for the  Digestion of Lacustrine
     Sediments for Subsequent Heavy Metal Analysis  by Atomic Absorption
     Spectrophotometry."  M.S. Thesis,  State University College of New
     York at Buffalo; Buffalo, New York.  77 p.  (1977).
16.  Anderson, J.  "A Study of the  Digestion of  Sediment by the HNOa-
     H2SOit and the HNOa-HCl Procedures."  Atomic  Absorption Newsletter
     13(1): 31-32 (197>0.
17-  Oliver, B. G.  "Heavy Metal Levels of Ottawa and Rideau River
     Sediments."  Environmental Science and Technology 7:135-137 (1973)
18.  Bothner, M. H., and Robertson, D.  E.  "Mercury Contamination of
     Sea Water Samples Stored in Polyethylene  Containers."  Anal.  Chem.
     ^7:592-595 (.1975).
19.  Lo, J.  M., and Wai, C. M.  "Mercury Loss  from  Water During Storage:
     Mechanisms and Preventions."  Anal. Chem. 1*7:1869-1870 (1975).
20.  Feldman, C.  "Preservation of  Dilute Mercury Solutions."  Anal.
     Chem. h6:99-102 (197*0.
21.  Agemian, H., and Chau, A. S. Y.  "A Method  for Determination of
     Mercury in Sediments by the Automated Cold  Vapor Atomic Absorption
     Technique After Digestion."  Analytica Chim. Acta 75:297-30*; (1975).
22.  Ure, A. M., and Shand, C. A.  "The Determination of Mercury in
     Soils and Related Materials by Cold-Vapor Atomic  Absorption
     Spectrometry."  Analytica Chim. Acta 72:63-77  (197*0 
23.  Martin, T. D., Kopp, J.  F., and Ediger, R.  D.   "Determining
     Selenium in Water, Wastewater, Sediment,  and Sludge by Flameless
     Atomic  Absorption Spectroscopy."  Atomic  Absorption Newsletter lU:
     109-116 (1975).
                                3-137

-------
                               NITROGEN
         (.Ammonia,  Nitrate,  Nitrite,  Total Kjeldahl, Organic)

            Nitrogen may be  distributed  among many  different forms  in
the environment.   These various nitrogen species  are important "because
they may contribute to eutrophication, they  may be  potentially toxic,
or they may affect the environmental  chemistry  of other constituents
such as metals by complexation and/or chelation.  However, the inter-
conversion of nitrogen species can create problems  during the analysis
of samples.  For example, nitrite can be chemically oxidized to nitrate
while algae and bacteria can alter ammonia,  nitrate, or organic nitrogen
concentrations.  In addition, poorly  protected  samples can pick up
ammonia from the atmosphere.  These effects  can be  minimized by
analyzing samples as soon as possible.
            This section presents analytical procedures for ammonia,
nitrate, nitrite, total Kjeldahl nitrogen, and  organic nitrogen.  Before
presenting these procedures, the relationship between these parameters
and their analytical techniques will  be  briefly summarized:
            a_.  Ammonia - This parameter is  measured either colori-
                          metrically  or  titrimetrically.  The analytical
                          procedures  measure total  ammonia.  Therefore,
                          if ammonia  toxicity is  of concern, temperature
                          and pH of the  original  sample should be
                          measured.
            b_.  Nitrate - One procedure  (Brucine  Sulfate) measures
                          nitrate directly.  The  remaining procedures
                          rely on a reduction of  nitrate to nitrite
                          and a subsequent quantification of the
                          nitrite. Thus, nitrate is calculated as
                          total nitrate  plus nitrite minus nitrite.
            c_.  Nitrite - The nitrate plus nitrite  procedures actually
                          measure nitrite.   Therefore, the nitrite
                          concentration  can  be  determined directly
                          by omitting the nitrate reduction step
                          (either the cadmium reduction column or
                          hydrazine sulfate  treatment).
            _d.  Total Kjeldahl Nitrogen  - The procedure catalytically
                          reduces organic nitrogen  to ammonia.  This
                          measurement, therefore, includes organic
                          nitrogen and ammonia.
                                3-138

-------
_e.  Organic Nitrogen - This parameter can be calculated as
              total Kjeldahl nitrogen minus ammonia nitrogen.
              Alternately, it may be determined by distilling
              off the ammonia at pH 9-5 and then running
              total Kjeldahl nitrogen on the sample residue.
                     3-139

-------
                                NITROGEN
                                (Ammonia)

                       Sample Handling and Storage

            A flow diagram summarizing the pertinent information regarding
sample handling and storage is presented in Figure 3-1.2.  Samples may be
collected in either glass or plastic.  They should be analyzed as. soon as
possible and preferably within 2k hr.  Sample stability can be improved
by adding sulfuric acid, tightly capping the sample bottle, and storing
at ^C until analyzed.  The volume of sample required will vary from 20
to 25 ml for the automated procedures to 500 ml for a manual procedure.

               Procedures for Water Samples (Wl, W2, S1A.)
                                             w.
Method 1:  Colorimetric, Automated Phenate1'2
            This procedure is suitable for samples with ammonia con-
centrations in the range of 0.01 to 2.0 mg Na-N/A.  It is based on the
reaction of ammonia with alkaline phenol and hypochlorite to form
indophenol.  The color is intensified with sodium nitroprusside and
measured colorimetrically.
Apparatus
Technicon AutoAnalyzer Unit (MI or Mil) consisting of:
      a^.  Sampler
      b_.  Manifold (Ml) or analytical cartridge (Mil)
      c_.  Proportioning pump
      d..  Heating bath with double delay coil (AAl)
      __.  Colorimeter equipped with 15-mm tubular flow cell and 630 to
          660-nm filters
      f_.  Recorder
      g_.  Digital printer for Mil (optional)
*
  References are on page 3-206.
                                  3-lUo

-------

CORE SAMPLE
4
*
WATER SAMPLE DREDGE SAMPLE CORE SECTION
J * 1

ACIDIFY FILTER N0 TRE-yENT STORE WET
1 1
STORE ACIDIFY
1
STORE


^^^^^^ F.IUTRIATF FRACTIONATE BIOA55AY STFAM DISTIIL

1 1 I 1
ANALYZE ANALYZE ANALYZE ANALYZE ANALYZE
(Wl) (W2) (SIA) (SIB) (SID)

1 SAMPLE DESIGNATION
H... 	 __.
H PURPOSE
CONTAINER
SAMPLE TREATMENT
PRESERVATIVE
Wl W2 W3 SIA SIB SIC SID
Total Water Soluble Used in Mobile Chemical Bioavail- Total
Cone. Water Elutriate Cone. Distribution ability Sediment
Cone. Cone.
G,P G,;P G,P G.P G,P G,P G,P
None Filter None None None None None
HaSOi, H2SO,, None 4C i|0C lt"C b'C
pH<2 pH<2 (Minimize Air Contact. Keep Field Moist.)

-------
Reagents

Distilled water:  special precaution must "be taken to ensure that
      distilled water is free of ammonia.  Such water is prepared by
      passage of distilled water through an ion exchange column comprised
      of a mixture of "both strongly acidic cation and strongly "basic
      anion exchange resins.  The regeneration of the ion exchange
      column should be carried out according to the instruction of the
      manufacturer.

NOTE:  All solutions must be made using ammonia- free water.

Sulfuric acid 5 N.:  air scrubber solution.  Carefully add 139 nil of cone.
      sulfuric acid to approximately 500 ml of ammonia- free distilled
      water .   Cool to room temperature and dilute to 1 H with ammonia-
      free distilled water.

Sodium phenolate:  using a 1-& Erlenmeyer flask, dissolve 83 g phenol in
      500 ml of distilled water.  In small increments, cautiously add,
      with agitation, 32 g of NaOH.  Periodically cool the flask under
      water faucet.  When cool, dilute to 1  with distilled water.
Sodium hypochlorite solution:  dilute 250 ml of a bleach solution con-
      taining 5-25 percent NaOCl (such as Clorox) to 500 ml with
      distilled water.  Available chlorine level should approximate
      2 to 3 percent.  Since Clorox is a proprietary product, its
      formulation is subject to change.  The analyst must remain alert
      to detecting any variation in this product significant to its use
      in this procedure.  Due to the instability of this product,
      storage over an extended period should be avoided.
Dissolve ethylenediamine-tetraacetate (EDTA) (5 percent):  dissolve
      50 g of EDTA (di sodium salt) and approximately six pellets of
      NaOH in 1 &'of distilled water.
NOTE:  On saltwater samples where EDTA solution does not prevent pre-
       cipitation of cations, sodium potassium tartrate solution may
       be used to advantage.  It is prepared as follows:
       Sodium potassium tartrate solution:  10 percent
             h H20.  To 900 ml of distilled water add 100 g sodium
             potassium tartrate.  Add two pellets of NaOH and a few
             boiling chips; boil gently for ^5 min.   Cover, cool, and
             dilute to 1 H with ammonia- free distilled water.  Adjust
             pH to 5.2 +_ 0.05 with H2SOu.  After allowing to settle
             overnight in a cool place, filter to remove precipitate.
             Then add 0.5 nil Brij-351* (available from Technicon Corpora-
             tion) solution and store in stoppered bottle.
Sodium nitroprusside (0.05 percent):  dissolve 0.5 g of sodium nitro-
      prusside in 1  of distilled water.
Stock solution:  dissolve 3.819 g of anhydrous ammonium chloride, NH^Cl,
      dried at 105C, in distilled water, and dilute to 1000 ml.  1.0 ml
      1.0 mg NH3-N.
                                   3-1^2

-------
Standard solution A:  dilute 10.0 ml of stock solution to 1000 ml with
      distilled water.  1.0 ml = 0.01 mg NH3-N.
Standard solution B:  dilute 10.0 ml of standard solution A to 100.0.ml
      with distilled water.  1.0 ml = 0.001 mg NH3-N.
Working ammonia standards should be prepared fresh on the day of use.
      They can be prepared by diluting either standard solution A or
      standard solution B as indicated below:
      NH3~N, mg/JL                          ml Standard Solution/100 ml
                                                   Solution B
         0.01                                          1.0
         0.02                                          2.0
         0.05                                          5-0
         0.10                                         10.0
                                                   Solution A
         0.20                                          2.0
         0.50                                          5.0
         0.80                                          8.0
         1.00                                         10.0
         1.50                                         15.0
         2.00                                         20.0
When freshwater samples are being analyzed, the working ammonia
      standards should be diluted to volume with ammonia-free distilled
      water.  When saltwater samples are being analyzed, the working
      ammonia standards should be diluted to volume with Substitute
      Ocean Water (SOW) that has the following composition:
                         Substitute Ocean Water  (SOW)
      Nad            2*1.53 g/&               NaHC03          0.20  g/
      MgCl             5-20 g/&               KBr             0.10  g/
      Na2S04           U.09 gA               H3B03           0.03  g/fc
      CaCl2            1.16 g/               SrCl            0.03  g/fc
      KC1              0.70 g/X,               NaF             0.003 g/A
If SOW is used, subtract its blank background response from the standards
      before preparing the standard curve.
Procedure
            Since the intensity of the color used to quantify the con-
centration is pH dependent, the acid concentration of the wash water and
the standard ammonia solutions should be approximately that of the samples.
For example, if the samples have been preserved with 2 ml cone. H2SOit/il,
the wash water and standards should also contain 2 ml cone.
                                3-1U3

-------
            For a working range of 0.01 to 2.00 mg NH3-N/& (AAl), set up
the manifold as shown in Figure 3-13..  For a working range of 0.01 to
1.0 mg NH3-N/ (Mil), set up the manifold as shown in Figure 3-1^.
Higher concentrations may be accommodated "by sample dilution.  Allow
both colorimeter and recorder to warm up for 30 min.  Obtain a stable
baseline with all reagents, feeding distilled water through sample line.
            For the MI system, sample at a rate of 20/hr, 1:1.  For the
Mil, use a 60/hr, 6:1 cam with a common wash..
            Arrange ammonia standards in sampler in order of decreasing
concentration of nitrogen.  Complete loading of sampler tray with unknown
samples.
            Switch sample from distilled water to sampler and begin
processing samples.
Calculations
            Prepare appropriate standard curve derived from processing
ammonia standards through manifold.  Compute concentration of samples by
comparing sample peak heights with standard curve.

Method 2:  Colorimetric, Automated 0-tolidine3
            This procedure is suitable for samples with ammonia con-
centrations in the range of 0.001 to 0.10 mg NH3-N/&.  It is based on
the reaction between ammonia and hypochlorite.   The product is reacted
with 0-tolidine and quantified colorimetrically.
Apparatus
Technicon AutoAnalyzer unit consisting of:
      ja.   Sampler
      b.   Manifold
      c_.   Proportioning pump
      d^.   Dialyzer
      _e.   Heating bath (UoC)
      f_.   Colorimeter equipped with a 50-mm flow cell and 420-nm filters
      g_.   Range expander
      h.   Recorder

-------
                    Figure 3-13.    AAI manifold for phenate determination of ammonia
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0.6 HYPOCHLORITE
0.6 NITROPRUSSIDE
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-------
                     Figure  3-1^.    AAII cartridge for phenate determination of ammonia
    SM = Smal1 Mixing Coil
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                               Scrubbed  through
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-------
Reagents
Alkaline complexing agent:

      Solution A:  dissolve 52 g sodium hydroxide, NaOH, in 1  deionized
                   water.

      Solution B:  dissolve Uo g sodium hexametaphosphate in 1  deio-
                   nized water.

      The alkaline complexing agent should be prepared fresh daily by
      mixing equal volumes of solution A and solution B (i.e. 100 ml A
      and 100 ml B).

Buffer:  dissolve 96 g hydrated disodium hydrogen phosphate and 10 g
      sodium dihydrogen phosphate in 5  deionized water (pH 7-5).
Sodium hypochlorite:  dilute sodium hypochlorite solution (Clorox is
      suitable) to approximately O.OOU percent available chlorine with
      deionized water.
Oxalic acid:  dissolve 20 g oxalic acid and 170 g monochloracetic acid
      in deionized water and make up to 1 .
Orthotolidine:  prepare by heating 1.2 g 0-tolidine dihydrochloride in
      120 ml cone, hydrochloric acid, HC1, at 60C for 1 hr; then
      adjust to a volume of 1  with distilled water.
Stock ammonia solution:  dissolve 3.819 g anhydrous ammonium chlorid'e,
      NH^Cl, dried at 100C, in distilled water and dilute to 1 9,.
      This solution contains 1000 mg/ N_.
Intermediate ammonia solution, 10 mg/:  dilute 10.00 ml stock ammonia
      solution to 1  with deionized water.
Standard ammonia solution, 1 mg/ N:  dilute 100 ml intermediate ammonia
      solution to 1  with deionized water.
Working ammonia standards:  using standard ammonia solution, prepare
      working ammonia standards in 100 ml-volumetric flasks.  Prepare
      fresh on the day of use.  The following standards are suggested:
      Ammonia E, mg/                ml Standard Solution/100 ml
           0.002                                 0.2
           0.01                                  1.0
           0.02                                  2.0
           0.05                                  5.0
           0.10                                 10.0
           0.15                                 15.0
           0.20                                 20.0

Procedure
            Arrange ammonia standards in sample tray in order of

decreasing nitrogen concentration.  Complete loading of tray with

samples.
                                 3-1^7

-------
NOTE:  Samples and standards may gain ammonia as the result of atmospheric
       contact.  One lab has found it convenient to cover loaded sample
       trays with Saran Wrap to prevent atmospheric contamination.  The
       protective covering is left in place during analysis and the
       metallic needlelike probe of the sampler is allowed to puncture
       the wrapping for each sample,3
            Samples and standards are run at a rate of 20/hr using a
manifold set up as shown in Figure 3-l5.
            If the sample response is off scale, one of the following
techniques must be implemented to analyze the samples:
            a.  Replace the colorimeter flow cell with a smaller flow cell.
            b.  Remove range expander.
            c_.  Replace sample line with a smaller line and add a
                distilled water line to make up the volume difference.
            d..  Dilute the samples prior to loading the sample tray.
Calculations
            Prepare a calibration curve derived from the peak heights
obtained with the standard solutions.
            Determine the concentration of ammonia in the samples by
comparing sample peak heights with the calibration curve.

Method 3:  Colorimetric or Titrimetric, Manual1'2'1*
            This method is applicable over a wide range of ammonia
concentrations.  Ammonia is distilled from the sample buffered at a pH
of 9-5 and collected in a boric acid solution.  The ammonia is then
quantified colorimetrically by nesslerization or titrimetrically.  The
former procedure is suitable for ammonia concentrations in the range of
0.05 to 1.0 mg NH3-N/Jl and the latter technique is useful for samples
with ammonia concentrations in the range of 1.0 to 25 mg NHa-N/^.
Apparatus
An all-glass distilling apparatus with an 800- to 1000-ml flask
Spectrophotometer or filter photometer for use at U25 nm and providing
      a light path of 1 cm or more
Nessler tubes:  matched Nessler tubes (APHA Standard) about 300 mm long,
      17 mm inside diameter, and marked at 225 mm +_ 1.5 mm inside
      measurement from bottom
                                  3-1^8

-------
                   Figure 3-15-     AA manifold for the 0-tolidine determination of ammmonia
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0.32 0-TOLIDINE
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                                                                                                       20/hr.

-------
Erlenmeyer flasks:  the distillate is collected in 500-ml glass-stoppered
      flasks.  These flasks should be marked at the 350- and the 500-ml
      volumes.  With such marking it is not necessary to transfer the
      distillate to volumetric flasks
Reagents

Distilled water should "be free of ammonia.  Such water is best prepared by
      passage through an ion exchange column containing a strongly acidic
      cation exchange resin mixed with a strongly "basic anion exchange
      resin.  Regeneration of the column should "be carried out according
      to the manufacturer's instructions.
NOTE:  All solutions must be made with ammonia-free water.
Ammonia chloride, stock solution:  1.0 ml = 1.0 mg NH3-N.  Dissolve
      3.819 g NHitCl in distilled water and bring to volume in a 1-Jl
      volumetric flask.

Ammonium chloride, standard solution:  1.0 ml = 0.01 mg.  Dilute 10.0 ml
      of stock ammonia chloride solution to 1 & in a volumetric flask.
Boric acid solution (20 g/):  dissolve 20 g H3B03 in distilled water and
      dilute to 1 H.

Mixed indicator:  mix two volumes of 0.2 percent methyl red in 95 percent
      ethyl alcohol with 1 volume of 0.2 percent methylene blue in
      95 percent ethyl alcohol.  This solution should be prepared fresh
      every 30 days.
NOTE:  Specially denatured ethyl alcohol conforming to Formula 3A or 30
       of the U. S. Bureau of Internal Revenue may be substituted for
       95 percent ethanol.
Nessler reagent:  dissolve 100 g of mercuric iodide and 70 g of potassium
      iodide in a small amount of water.  Add this mixture slowly, with
      stirring, to a cooled solution of 160 g of NaOH in 500 ml of water.
      Dilute the mixture to 1 I.  If this reagent is stored in a Pyrex
      bottle out of direct sunlight, it will remain stable for a period
      of up to 1 year.
NOTE:  This reagent should give the characteristic color with ammonia
       within 10 min after addition and should not produce a precipi-
       tate with small amounts of ammonia (0.0^ mg in a 50-ml volume).
Borate buffer:  add 88 ml of 0.1 N NaOH solution to 500 ml of 0.025 M
      sodium tetraborate solution (5-0 g anhydrous NaaBijO? or 0.5 g
      Na2Bit07  10H20 per liter) and dilute to 1 .
Sulfuric acid, standard solution:  0.02 N, 1 ml = 0.28 mg NH3-N.  Prepare
      a stock solution of approximately 0.1 N_ acid by diluting 3 ml of
      cone. HaSOit (sp.  gr. 1.8U) to 1 I with C02-free distilled water.
      Dilute 200 ml of this solution to 1 & with COa-free distilled water.

NOTE:  An alternate and perhaps preferable method is to standardize the
       approximately 0.1 N_ HaSOif solution against a 0.100 N_ Na2C03
       solution.  By proper dilution, the 0.02 N_ acid can then be prepared.

                                  3-150

-------
       Standardize the approximately 0.02 N acid against 0.0200 N
       Na2 C03  solution.   This last solution is prepared by dissolving
       1.060g anhydrous Na^ C03 ,  oven dried at 1^0C, and diluting to
       1000 ml with C02-free distilled water.
Sodium hydroxide, 1 N_:  dissolve Uo g NaOH in ammonia-free water and
      dilute to 1 H.
Dechlorinating reagents:  a number of dechlorinating reagents may be
      used to remove residual chlorine prior to distillation.  These
      include:
      a..  Sodium thiosulfate (0.01U2 N):   dissolve 3.5 g NazSaOs 
          5H20 in distilled water and dilute to 1 .  One milliliter
          of this solution will remove 1 mg/&  of residual chlorine
          in 500 ml of sample.
      b_.  Sodium arsenite (0.01^2 IT):   dissolve 1.0 g NaAsOa  in
          distilled water and dilute to 1 .
Procedure
            Preparation of equipment:   add 500 ml of distilled water
to an 800-ml Kjeldahl flask.  The addition of boiling chips which have
been previously treated with dilute NaOH will prevent bumping.  Steam
out the distillation apparatus until the distillate shows no trace of
ammonia with Nessler reagent.
            Sample preparation:   remove the residual chlorine in the
sample by adding dechlorinating agent equivalent to the chlorine
residual.  To 1*00 ml of sample,  add 1 IT NaOH until the pH is 9.5,
checking the pH during addition with a pH meter or by use of a short-
range pH paper.
            Distillation:  transfer the sample, the pH of which has
been adjusted to 9.5, to an 800-ml Kjeldahl flask and add 25 ml of
borate buffer.  Distill 300 ml at the rate of 6 to 10 ml/min into 50 ml
of 2 percent boric acid contained in a 500-ml Erlenmeyer flask.
NOTE:  The condenser tip or an extension of the condenser tip must extend
       below the level of the boric acid solution.
            Dilute the distillate to 500 ml with distilled water and
nesslerize an aliquot to obtain an approximate value of the ammonia-
nitrogen concentration.   For concentrations above 1 mg/Jl, the ammonia
should be determined titrimetrically.   For concentrations below this
value, it is determined colorimetrically.
                                  3-151

-------
            Titrimetric determination:  add 3 drops of the mixed indica-
tor to the distillate and titrate the ammonia with the 0.02 1J HgSO^,
matching the end point against a blank containing the same volume of
distilled water and HaBOa solution.
            Colorimetric determination;  prepare a series of Nessler
tube standards as follows:
          ml of Standard
      1.0 ml = 0.01 rag MH3-H                    mg NHa-N/50.0 ml
                0.0                                    0.0
                0.5                                    0.005
                1.0                                    0.01
                2.0                                    0.02
                3.0                                    0.03
                U.O                                    O.OU
                5.0                                    0.05
                8.0                                    0.08
               10.0                                    0.10
Dilute each tube to 50 ml with distilled water; add 2.0 ml of Nessler
reagent and mix.  After 20 min, read the absorbance at 425 nm against
the blank.  From the values obtained, plot absorbance vs. mg NH3-N for
the standard curve.  Determine the ammonia in the distillate by
nesslerizing 50 ml or an aliquot diluted to 50 ml and reading the
absorbance at ^25 nm as described above for the standards.  Ammonia-
nitrogen content is read from the standard curve.
            It is not imperative that all standards be distilled in the
same manner as the samples.  It is recommended that at least two
standards (a high and low) be distilled and compared to similar values
on the curve to ensure that the distillation technique is reliable.
If distilled standards do not agree with undistilled standards, the
operator should find the cause of the apparent error before proceeding.
Calculations
            Titrimetric:
                       mg/Jl M3.N = A * 0.28 x 1000
                                           o
where
     A = ml 0.02 N H2SOit used
     S = ml sample
                                 3-152

-------
            Spectrophotometric:
                           , .T1I     A x 1000   B
                        mg/A NH3-N =	^	x --
where
     A = mg NHa-N read from standard curve
     B = ml total distillate collected, including boric acid and dilution
     C = ml distillate taken for nesslerization
     D = ml of original sample taken
                                 3-153

-------
                  Procedures for Sediment Samples (SID)

            The determination of ammonia in sediments should- be considered
operationally defined because of the subjective nature of sample pre-
treatment.  Ammonia is extracted from sediments using a salt solution.
While this will remove the exchangeable ammonia, similar to the cation
exchange capacity procedure, and is considered a reproducible procedure,5
the samples are not subjected to exhaustive digestion.  However, the use
of acid may destroy nitrites in the sample and acid digestion may result
in the conversion of organic nitrogen to ammonia.

                       Sample Handling and Storage

            It is recommended that ammonia determinations only be run
on sediment samples stored in a field moist condition (Figure 3-12).
This recommendation is based on the possibility that ammonia may be lost
by volatilization during the drying or thawing of samples stored in
other conditions.  In addition, samples should be processed in a week
or less to minimize the effects of ammonia conversion or ammonia
absorption.
Method 1:  Potassium Chloride Extraction
Apparatus
Wrist-action or equivalent mechanical shaker
Filtration apparatus
Erlenmeyer flasks, 150 ml
Volumetric flasks, 100 ml
Reagents
Ammonia- free distilled water
Potassium chloride, 2 M:   dissolve 1^9-H , KC1 in ammonia-free distilled
      water and dilute to 1 H.
Procedure
            Weigh a 20-g  sample of wet sediment and transfer to a 150-ml
Erlenmeyer flask.  Add 50 ml of 2 M KC1 and seal the flask.
            Shake the sample on a wrist-action or equivalent mechanical

-------
shaker for 30 min.  Since the procedure is operationally defined, the
shaking time should be standardized for all samples.
            Filter the sample through a prewashed 0.^5-ym pore-size
membrane filter.  Collect the filtrate in a 100-ml volumetric flask.
Wash the solids with a second 50-ml portion of 2 M KC1.  Repeat the
filtration process and add the filtrate to the volumetric flask.  Dilute
the sample to volume with ammonia-free distilled water.
            Analyze the samples by one of the procedures listed for
ammonia in water.
Calculations
            Determine the ammonia concentration of the KC1 leachate
using the appropriate standard curve.  Calculate the ammonia concentra-
tion of the sediment sample as follows:
                      M   h   i  4- -u   \   (x)(y)(lOOQ)
              Ammonia-N mg/kg (wet basis) = v [/ w-'	
                                                  g
where
     x = ammonia concentration in leachate, mg/&
     y = sample volume, & (0.1 & as described)
     g = wet weight of sediment sample extracted, g
            Ammonia concentrations on a dry weight basis can be
calculated by dividing the wet weight concentration by the percent
solids in the sediment sample, expressed as a decimal fraction.

Method 2:  Distillation
            Ammonia is distilled from a sample and trapped in a boric
acid solution.  The distillate is then analyzed using one of the ammonia
methods listed in the section for water analysis.  The apparatus and
reagents will depend on the method selected.
Apparatus
Kjeldahl digestion apparatus
Reagents
Ammonia-free water.
Phosphate buffer solution, pH 7.^:  dissolve 1^.3 g anhydrous potassium
      dihydrogen phosphate, KHaPOt,, and 68.8 g anhydrous dipotassium
      hydrogen phosphate, KaHPOi*, and dilute to 1 & with ammonia-free
      water.
                                  3-155

-------
Boric acid solution:  dissolve 20 g anhydrous boric acid,  H3B03 ,  in
      ammonia-free water and dilute to 1  .
Procedure
            Weigh a 0.5- "to 1.0-g sample of  wet sediment.   Transfer the
sample to a 100-ml Erlenmeyer flask and add  approximately  50 ml ammonia-
free water and 3 to U drops concentrated sulfuric acid.  This will sta-
bilize the ammonia and the procedure can be  interrupted  at this point
if necessary.
            Steam out the distillation apparatus.  Add 500 ml ammonia-
free water, 10 ml phosphate buffer, and a few boiling stones to an 800-ml
flask and steam the apparatus until there is no trace of ammonia in the
distillate.
            Transfer the acidified sediment  slurry to an 800-ml Kjeldahl
flask and add 500 ml ammonia-free water and  a few boiling  stones.   Boil
for a few minutes to remove any sulfides that may be present.  This step
will also remove any volatile organics such  as formaldehyde that may
interfere with the nesslerization procedure.
NOTE:  Sulfide interferences may also be removed by precipitating the
       sulfide with lead carbonate.
            Neutralize the sample to a pH of about 6.6 and add 10 ml
phosphate buffer.  Distill over 300 ml of sample at a rate of 6 to 10 ml/
min and collect in 50 ml boric acid solution.  Dilute the  distillate to
500 ml with ammonia-free water.
            Analyze the distillate using either the automated methods,
direct nesslerization, or titration as described earlier.   Nesslerization
should be used if the ammonia concentration  is less than 1 mg NH3-N/,
and titration should be used when the ammonia concentration is greater
than 1 mg MH3-N/JL
            The residue in the distillation  flask can be used to deter-
mine organic nitrogen.  By subjecting the residue to a Kjeldahl diges-
tion, the result is a measure of organic carbon since this parameter is
defined as total Kjeldahl nitrogen minus ammonia nitrogen.
Calculations
        Ammonia-nitrogen mg NH3-N/kg (wet basis) = ^
                                 3-156

-------
where
     x = ammonia concentration in distillate, mg/&
     y = volume of distillate, 
     g = wet weight of sediment used, g

        Ammonia-nitrogen mg NH3-N/kg (dry basis) =

                                                     g (7 o)
where
       x = ammonia concentration in distillate, mg/&
       y = volume of distillate, 
       g = wet weight of sediment used, g
     % S = percent solids as a decimal fraction

Method 3:  Distilled Water Extraction
            Ammonia is separated from the sediment using an aqueous
extraction technique.  The liquid phase is then analyzed for ammonia
using the method of choice in the Procedures for Water Samples section,
which begins on page 3-1^0.  This procedure is operationally defined
and, therefore, must be carefully followed.
Procedure
            Weigh out 0.5 to 1.0 g wet sediment and transfer to a
250-ml Erlenmeyer flask.  Add 100 ml ammonia- free distilled water.
Thoroughly mix and allow suspension to settle overnight.
            Transfer to a centrifuge tube and centrifuge 10 min at
2000 rpm.  Decant liquid to a 200-ml volumetric flask.
            Add 50 ml ammonia-free distilled water to the sediment  and
mix.  Centrifuge as before and add the wash to the initial extract.
            Dilute the combined extract to volume with ammonia-free
distilled water and analyze by the ammonia method of choice in the
"Procedures for Water Samples" section.
Calculations
            The ammonia concentration of the sediment samples is
calculated as follows:
                Ammonia mg/kg (wet weight) = -(x) (y) (1000)
                                  3-157

-------
                Ammonia mg/kg (dry weight)  = U
where
       x = ammonia concentration in extract,  mg/&
       y = volume of extract, & (0.2 as written)
       g = wet weight of sediment ,  g
     % S = percent solids in sediment (as decimal fraction)
                                  3-158

-------
                                NITROGEN
                                (Nitrate)

                      Sample Handling and  Storage

            Samples may be collected and stored in either plastic  or  glass
containers.  The accepted preservative for this parameter is sulfuric
acid to a pH of 2 and refrigeration at ^C.   This combination of preserva-
tives may stabilize samples for as long as U weeks  but it is generally
recommended that samples be analyzed within 2h hr.  This information  is
presented in Figure 3-16.
            The volume of sample required  depends on the analytical
method of choice and ranges from 10 ml for the nitrate-specific  procedure
(Brucine Sulfate) to 100 ml for the manual nitrate-nitrite procedure.

               Procedures for Water Samples (Wl, W2, S1A)

Method 1;  Colorimetric, Manual, Brucine Sulfate1'2
            The procedure is based on the  reaction between nitrate and
brucine sulfate in a highly acidic medium.  The resulting product  is
then quantified colorimetrically.  Temperature control is a critical
aspect of this procedure.  The method is applicable to samples with
concentrations of 0.1 to 2.0 mg N03-N/&.
Apparatus
Spectrophotometer or filter photometer suitable for measuring absorbance
      at UlO nm
Sufficient number of kO- to 50-ml glass sample tubes for reagent blanks,
      standards, and samples
Neoprene-coated wire racks to hold sample  tubes
Water bath suitable for use at 100C.  This bath should contain  a  stirring
      mechanism so that all tubes are at the same temperature and  should
      be of sufficient capacity to accept  the required number of tubes
      without significant drop in temperature when the tubes are immersed
Water bath suitable for use at 10 to 15C
Reagents
Distilled water free of nitrite and nitrate is to be used in preparation

                                  3-159

-------
                 Figure 3-l6.     Handling  and storage of samples for nitrate analysis

CORE SAMPLE
*

WATER SAMPLE )REDGE SAMPLE CORE SECTION
i * 1
4 * * *
ACIDIFY FILTER N0 T^jTMENT STORE WET
I 1
STORE ACIDf.FY
1
STORE
< '
+ 4
. _* FIIITRIATF HlflASSAY
" . (SIC)
i I
ANALYZE ANALYZE ANALYZE ANALYZE
U) (Wl) (W2) (SIA) (SID)
H
 SAMPLE DESIGNATION
PURPOSE
CONTAINER
SAMPLE TREATMENT
PRESERVATIVE
STORAGE TIME
DIGESTION SOLUTION
Wl W2 W3 SIA SIC SID
Total Water Soluble Used in Mobile Bioavall- Total
Cone. Water Elutriate Cone. ability Sediment
Cone. Cone.
G,P G,P G.P G.P G,P G,P
None Filter None None None None
H2SOi, H2SOi, None I( C ^ C 4 C
pH<2 pH<2 (Minimize Air Contact. Keep Field Moist.)
ltC !,<>C
2k hr 2>t hr <1 w <1 w <1 w <1 w
Distilled
Water
SAMPLE VOLUME OR WEIGHT
                      20-100 ml    20-100 ml
                                                                                   5-10 g

-------
      of all reagents and standards.
Sodium chloride solution (30 percent):  dissolve 300 g Nad in distilled
      water and dilute to 1 .
Sulfuric acid solution:  carefully add 500 ml cone. Ha S0i to 125 ml
      distilled water.  Cool and keep tightly stoppered to prevent
      absorption of atmospheric moisture.
Brucine-sulfanilic acid reagent:  dissolve 1 g Brucine Sulfate
       (Cg^gNaO,, )2  ' f^SO,, ' ?H20  and 0.1 g sulfanilic acid
      (NH2C6H4S03H  H20) in TO ml hot distilled water.  Add
      3 ml cone. HC1, cool, mix, and dilute to 100 ml with distilled
      water.  Store in a dark bottle at 5C.  This solution is stable
      for several months; the pink color that develops slowly does
      not affect its usefulness.  Mark bottle with warning:  CAUTION:
      Brucine Sulfate is toxic; take care to avoid ingestion.
Potassium nitrate stock solution:  1.0 ml = 0.1 mg N03-N.  Dissolve
      0.7218 g anhydrous potassium nitrate, KN03, in distilled water
      and dilute to 1 & in a volumetric flask.  Preserve with 2 ml
      chloroform per liter.  This solution is stable for at least
      6 months.
Potassium nitrate standard solution:  1.0 ml = 0.001 mg N03-W.  Dilute
      10.0 ml of the stock solution to 1 & in a volumetric flask.  This
      standard solution should be prepared fresh weekly.
Acetic acid (.1 + 3):  dilute 1 volume glacial acetic acid (CH3COOH) with
      3 volumes of distilled water.
Sodium hydroxide (l N_):  dissolve ko g of NaOH in distilled water.  Cool
      and dilute to 1 .
Procedure
            Adjust the pH of the samples to approximately 7 with acetic
acid or sodium hydroxide.  If necessary, filter to remove turbidity.
            Set up the required number of sample tubes in the rack to
handle reagent blank, standards, and samples.  Space tubes evenly
throughout the rack to allow for even flow of bath water between the
tubes.  This should assist in achieving uniform heating of all tubes.
            Pipet 10.0 ml of standards and samples or an aliquot of
the samples diluted to 10.0 ml into the sample tubes.
            Pipet 10.0 ml sulfuric acid solution into each tube and
mix by swirling.  Place samples and standards in a cold water bath
(.0 to 10C) and do not continue until all samples have reached tempera-
ture equilibrium.
            Add 0.5 ml brucine-sulfanilic acid reagent to each tube and

                                  3-l6l

-------
carefully mix by swirling.   Place samples in a 100C water bath  for
exactly 25 min.

            Remove rack of  tubes from the hot water bath  and  immerse  in
the cold water bath and allow to reach thermal equilibrium (20  to 25C)

            Read absorbance against the reagent blank  at  1*10  run  using
a 1-cm or longer cell.

            CAUTION:   The procedure is sensitive to temperature,  ionic
strength, and color effects of interferences.   The following  procedures
should be followed when appropriate:
            a_.  Immersion of the tube rack into the bath  should  not
                decrease the temperature of the bath more than 1 to  2C.
                In order to keep this temperature decrease to an
                absolute minimum, flow of bath water between  the
                tubes should not be restricted by crowding too many
                tubes into  the rack.   If color development in the
                standards reveals discrepancies in the procedure,
                the operator should repeat the procedure  after
                reviewing the temperature control steps.
            b_.  If samples  are saline, the ionic strength is  buffered
                prior to acidification of the samples.  Add 2 ml
                30 percent  sodium chloride solution to the reagent
                blank,  standards, and samples.  Mix by swirling  and
                place samples in the cold water bath (0  to 10C).
                This step is not necessary for freshwater samples.
            .  Samples that are colored or contain dissolved organic
                matter  that can cause the sample to become colored on
                heating must be run in duplicate. These  samples  are
                colorimetric blanks and should receive all reagent
                additions except brucine-sulfanilic acid.  They  should
                receive 0.5 ml distilled water to compensate  for
                dilution effects.
Calculations
            Obtain a standard curve by plotting the absorbance of

standards run by the above  procedure against mg N03-N/.  (The color
reaction does not always follow Beer's law.)  Subtract the absorbance
of the sample without the brucine-sulfanilic reagent from the absorbance
of the sample containing brucine-sulfanilic acid and determine mg NO

Multiply by an appropriate  dilution factor if less than 10 ml of sample

is taken.
                                 3-162

-------
Method 2:  Colorimetric, Automated, Cadmium Reduction
            The procedure is applicable to the quantification of either
nitrite singly or nitrate plus nitrite combined.   Nitrate is reduced in a
cadmium reduction column and the total nitrite (original nitrite plus
reduced nitrate) is diazotized and coupled with naphthyl-ethylenediamine
dihydrochloride.  The azo dye that is formed is then quantified colori-
metrically.
            Nitrite can be determined separately by omitting the cadmium
reduction step.  Nitrate can then be calculated by running the sample a
second time using the cadmium reduction column and subtracting the nitrite
concentration from the combined nitrate-nitrite concentration.
            It is necessary to continually monitor the performance  of
the reduction column.  This is accomplished by running nitrate and
nitrite standards of equal concentration.   When discrepancies occur
(nitrate standard less than nitrite standard), the column is not
operating efficiently and must be replaced.
            Samples to be analyzed for nitrate or nitrate plus nitrite
must not be preserved with mercuric chloride.
Apparatus
Technicon AutoAnalyzer (.AAI or Mil) consisting of the following
      components:
      a_.  Sampler
      b_.  Manifold (AAl) or analytical cartridge (AAIl)
      c_.  Proportioning pump
      d_.  Colorimeter equipped with a 15- or 50-mm tubular flow cell and
          5^0-nm filters
      j;.  Recorder
      f_.  Digital printer for AAII (optional)
Reagents
Granulated cadmium:  hO-60 mesh (E. M. Laboratories, Inc., 500 Executive
      Boulevard, Elmsford, New York 10523; Cat. 2001 Cadmium, Coarse
      Powder).
Copper-cadmium:  the cadmium granules (new or used) are cleaned with
      dilute HC1 and copperized with 2 percent solution of copper
      sulfate in the following manner:
      a_.  Wash, the cadmium with HC1 and rinse with distilled water.
          The color of the cadmium so treated should be silver.
                                 3-163

-------
      b_.  Swirl 10 g cadmium in 100-ml portions of 2 percent solution
          of copper sulfate for 5 niin or until blue color partially
          fades; decant and repeat with fresh copper sulfate until a
          brown colloidal precipitate forms.

      c_.  Wash the cadmium-copper with distilled water  (at least  10
          times) to remove all the precipitated copper.   The color of
          the cadmium so treated should be black.
Preparation of reduction column MI:   the reduction column is an  8- by
      50-mm glass tube with the ends  reduced  in diameter to permit
      insertion into the system.  Copper cadmium granules are placed
      in the column between glass wool plugs.   The packed reduction
      column is placed in an upflow 20-deg incline to minimize chan-
      neling.  See Figure 3-17.

Preparation of reduction column Mil:  the reduction column is a  U-
      shaped, 35-cm-long, 2-mm-I.D. glass tube (see NOTE).  Fill  the
      reduction column with distilled water to prevent  entrapment of
      air bubbles during the filling operations.  Transfer the
      copper-cadmium granules to the reduction column and place a
      glass wool plug in each end.  To prevent entrapment of air
      bubbles in the reduction column, be sure that all  pump tubes
      are filled with reagents before putting the column into the
      analytical system.
NOTE:  A 0.08l-in.-I.D. pump tube (purple) can be used  in place of
       the 2-mm glass tube.
Distilled water:  because of possible contamination, this should  be
      prepared by passage through an ion exchange column comprised of
      a mixture of both strongly acidic-cation and strongly basic-
      anion exchange resins.  The regeneration of the ion exchange
      column should be carried out according  to the manufacturer's
      instructions.
Color reagent:  to approximately 800  ml of distilled water, add,  while
      stirring, 100 ml cone, phosphoric acid,  ko g sulfanilimide, and
      2 g N-1-naphthyl-ethylenediamine dihydrochloride.   Stir until
      dissolved and dilute to 1 A.  Store in  brown bottle and keep in
      the dark when not in use.  This solution is stable for several
      months.
Dilute hydrochloric acid, 6 N_:  dilute 50 ml  of cone. HC1 to 100  ml
      with distilled water.
Copper sulfate solution, 2 percent:  dissolve 20 g of CuSOi,  5H20 in
      500 ml of distilled water and dilute to 1 .
Wash solution:  use distilled water for unpreserved samples.   For
      samples preserved with H2SOit, use 2 ml  H2SOi, per  liter of wash
      water.

Ammonium chloride-EDTA solution:  dissolve 85 g of reagent grade
      ammonium chloride and 0.1 g of disodium ethylenediamine
                                3-161*

-------
          Figure 3-17.    Copper cadmium reduction column
                                           GLASS WOOL
INDENTATIONS  FOR  SUPPORTING
         CATALYST
         FLOW
                     Cd - TURNINGS
                        TILT COLUMN TO 20 POSITION
                               3-165

-------
      tetracetate in 900 ml of distilled water.   Adjust the pH to 8.5
      with cone,  ammonium hydroxide and dilute to 1 &.   Add 0.5 ml
      Brij-35 (available from Technicon Corporation) .
Stock nitrate solution:   dissolve T-218 g KKOs and dilute to 1  in a
      volumetric  flask with distilled water.   Preserve  with 2 ml of
      chloroform per liter.  Solution is stable for 6 months.  1 ml =
      1.0 mg NOa-N.
Stock nitrite solution:   dissolve 6. 072 g KNOa in 500 ml of distilled
      water and dilute to 1  in a volumetric flask. Preserve with
      2 ml of chloroform and keep under refrigeration.   1.0 ml = 1.0 mg
      NOa-N.

Standard nitrate solution:  dilute 10.0 ml of stock nitrate solution
      to 1000 ml.  1.0 ml = 0.01 mg NOa-N.  Preserve with 2 ml of
      chloroform per liter.  Solution is stable for 6 months.
Standard nitrite solution:  dilute 10.0 ml of stock nitrite solution to
      1000 ml.  1.0 ml = 0.01 mg NOz-N.  Solution is unstable; prepare
      as required.
Using standard nitrate solution, prepare the following  standards in
      100-ml volumetric flasks.  At least one nitrite  standard should
      be compared to a nitrate standard at the same concentration to
      verify the efficiency of the reduction column.

      Cone, mg NOa-N or NQa-N        ml Standard Solution/100 ml
               0.00                             0
               0.05                             0.5
               0.10                             1.0
               0.20                             2.0
               0.50                             5.0
               1.00                            10.0
               2.00                            20.0
               U.OO                            1*0.0
               6.00                            60.0

NOTE:  When the samples to be analyzed are saline waters, Substitute
       Ocean Water (SOW) should be used for preparing the standards;
       otherwise, distilled water is used.  A tabulation of SOW
       composition follows:
       NaCl  - 2^.53  g/      MgCl2 - 5-20

       CaC3.2 -  1.16  g/      KC1   - 0.70 g/       NaHC03 - 0.20

       KBr   -  0.10  g/Jl      H3B03 - 0.03 g/       SrCl2  - 0.03 g/S,

       NaF   -  0.003 g/

Procedure

            Adjust the sample pH between 5 and 9 using either cone .

HC1 or cone. NH^OH.

            Set up the manifold as shown in Figure 3-18 for an AAI
                                 3-166

-------
Figure 3-18.     AAI manifold for nitrate determination following cadmium reduction

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-------
or Figure 3-19 for an Mil.   Take care not to introduce air  into  the
reduction column of the Mil during set up and initial operation.   If
the MI option is selected,  "be sure to incline the reductant column
at approximately 20 deg with flow from bottom to top.
            Allow "both colorimeter and recorder to warm up for  30 min.
Obtain a stable baseline with all reagents,  feeding distilled water
through the sample line.
NOTE:  Condition column by running 1 mg/& nitrate standard for  10 min
       if a new reduction column is being used.  Subsequently wash
       the column with reagents for 20 min.
            Place appropriate nitrate and/or nitrite standards  in
sampler in order of decreasing concentration of nitrogen.  Complete
loading of sampler tray with unknown samples.  For the MI system,
sample at a rate of 30/hr, 1:1 cam and a common wash.   For the  Mil,
use a itO/hr, U:l cam and a common wash.
            Switch sample line to sampler and start analysis.
Calculations
            Prepare appropriate standard curve or curves derived  from
processing N02 and/or NOs standards through manifold.   Compute  concen-
tration of samples by comparing sample peak heights with standard curve.

Method 3:  Colorimetric, Automated, Hydrazine Reduction
            The only major difference between this method  and Method  2,
Cadmium Reduction, is the reducing agent used to reduce nitrate to
nitrite.  Hydrazine sulfate is used in place of the cadmium  reduction
column with this procedure.   The total nitrite (original nitrite  plus
reduced nitrate) is then diazotized and determined colorimetrically as
before.  The method is applicable to a combined nitrate-nitrite
concentration of 0.01 to 10 mg N/JL
            Nitrite can be determined separately by omitting the  use
of hydrazine sulfate.  Nitrate can then be calculated by subtracting
the nitrite concentration from the combined nitrate-nitrite  concentration.
Apparatus
Sampler
Manifold MI or Mil
                                3-168

-------
          Figure 3-19.     AAII cartridge for the determination of nitrate following cadmium reduction
    SM = Small  Mixing Coil
    LM = Large  Mixing Coil
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                                                0. "?2   AIR
                                                1.2
AMMONIUM CHLORIDE
                                                                        0.32   SAMPLE
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                                                                        0.32   COLOR REAGENT
                                                0.6    WASTE
                                                                        1.0    WASTE
                                                                        2.0    WASH

-------
Proportioning pump

Heating bath 32 C AAI or 37 C Mil

Continuous filter

Colorimeter equipped with an 8-, 15-, or 50- mm flow cell and 529-nm
      filters

Reagents

Color development reagent:  to approximately 500 ml of distilled water
      add 200 ml concentrated phosphoric acid (sp. gr . 1.8sM 10 g
      sulfanilamide (MCelUSOaNHz ) , followed by 0.8 g N (l-Naphthyl)
      ethyl en ediamine dihydrochloride.  Dilute the solution to 1 & with
      distilled water and store in a dark bottle in the refrigerator.
      This solution is stable for approximately 1 month.

Copper sulfate stock solution:  dissolve 2.5 g of copper sulfate
               5H20) in distilled water and dilute to 1 &.
Copper sulfate dilute solution:  dilute 20 ml of stock solution to 2 H
      with distilled water.

Sodium hydroxide stock solution (10 N_) :   dissolve ^00 g NaOH in 750 ml
      distilled water, cool, and dilute to 1 &.

Sodium hydroxide (.1.0 H_) :   dilute 100 ml of stock NaOH solution to 1 & .

Hydrazine sulfate stock solution:  dissolve 27-5 g of hydrazine
      sulfate (Mi*  HaSOtf) in 900 ml of distilled water and dilute
      to 1 i .  This solution is stable for approximately 6 months.
      Mark container with appropriate warning:   CAUTION:  Toxic if
      ingested.

Hydrazine sulfate dilute solution:

      a_.  When using an AAI, dilute 55 ml stock hydrazine sulfate to
          1 & with distilled water.

      b_.  When using an AAII, dilute 22 ml stock hydrazine sulfate to
          1 H with distilled water.

Stock nitrate solution (.100 mg/ N03-N) :   dissolve 0.7218 of KN03,
      oven dried at 100 to 105 C for 2 hr, in distilled water and
      dilute to 1 H.  Add 1 ml chloroform as a preservative.  Solution
      is stable for approximately 6 months.  1 ml = 0.1 mg N.

Stock nitrite solution (100 mg/ NOa-N):   dissolve 0.6072 g KW02 in
      500 ml of distilled water and dilute to 1 &.  Preserve with 2 ml
      of chloroform and keep under refrigeration.  1 ml = 0.1 mg N.

Standard nitrate solution:  dilute 100 ml of stock nitrate solution
      to 1 .  1 ml = 0.01 mg N.

Using the stock nitrate solution, prepare the standards on the
      following page in 100-ml volumetric flasks.  At least one
      nitrite standard should be compared to a nitrate standard at
      the concentration to verify the efficiency of the reduction.
                                3-170

-------
     ml of Stock Solution/100 ml              Cone., mg N03-N/&
                 0.5                                0.5
                 1.0                                1.0
                 2.0                                2.0
                 3.0                                3.0
                 h.O                                k.O
                 5.0                                5.0
                 8.0                                8.0
                10.0                               10.0
Procedure
            Set up the appropriate manifold as suggested  in  either
Figure 3-20 (AMI) or Figure 3-21 (AAl).   Both procedures require the
use of a continuous filter to remove precipitate that will interfere
with the colorimetric procedure.
            Allow both colorimeter and  recorder to warm up for 30 min.
Obtain a stable baseline with all reagents, feeding distilled water
through the sample line.
            Run a 2.0-mg/ NOs-N and a  2.0-mg/ N02-N standard through
the system to check for 100 percent reduction of nitrate  to  nitrite.
The two peaks should be of equal height.   If they are not, the con-
centration of the hydrazine sulfate solution must be adjusted as
follows.  If the NOa peak is lower than that of the NOa peak, the
concentration of hydrazine sulfate should be increased until they are
equal.  If the NOs peak, is higher than  the nitrite, the concentration
of the hydrazine sulfate should be reduced.   When the correct concen-
tration of hydrazine sulfate has been determined, no further adjust-
ment should be necessary.
            Place standards in the sample tray in order of decreasing
concentration.  Complete loading of tray  with samples.
            Process samples at the rate of 30/hr.
Calculations,
            Prepare a standard curve by plotting peak heights of
processed standards against known concentrations.  Compute concentra-
tions of samples by comparing sample peak heights with the standard
curve.
                                 3-171

-------
          Figure  3-20.     AAI manifold for the determination of nitrate following hydrozine reduction
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-------
            Figure 3-21.    Mil cartridge for the determination of nitrate following hydrazine reduction
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-------
Method U:  Colorimetric, Manual, Cadmium Reduction7

            This procedure is similar to Method 2 except that  the
samples are processed manually rather than automatically.  The same uses
and cautions, therefore, apply to this method.

            Samples to be analyzed for nitrate  or nitrate plus nitrite
must not be preserved with mercuric chloride as it poisons the cadmium
reduction column.

Apparatus

Reduction column:  the column in Figure 3-22 was constructed from a
      100-ml pipet by removing the top portion.  This  column may also
      be constructed from two pieces of tubing  joined  end to end.
      A 10-mm length of 3-cm-I.D.  tubing is joined to  a  25-cm  length
      of 3.5-mm-I.D. tubing
Spectrophotometer for use at 5^0 nm, providing  a light path of 1 cm or
      longer

Reagents
Granulated cadmium:  kd to 60 mesh (E.  M.  Laboratories,  Inc.,  500
      Executive Boulevard, Elmsford, New York 10523; Cat.  2001
      Cadmium, Coarse Powder).
Copper-Cadmium:  the cadmium granules (new or used) are  cleaned with
      dilute HC1 and copperized with 2 percent  solution  of copper
      sulfate in the following manner:
      a_.  Wash the cadmium with dilute HC1 and  rinse with distilled
          water.  The color of the cadmium should be silver.
      b_.  Swirl 25 g cadmium in 100-ml portions of a 2 percent
          solution of copper sulfate for 5 uiin  or until  blue color
          partially fades; decant and repeat with fresh  copper sulfate
          until a brown colloidal precipitate forms.
      c_.  Wash the copper-cadmium with distilled water (at least 10
          times) to remove all the precipitated copper.   The color of
          the cadmium so treated should be black.
Preparation of reaction column:  insert a glass wool plug into the
      bottom of the reduction column and fill with distilled water.
      Add sufficient copper-cadmium granules to produce  a column
      18.5 cm in length.  Maintain a level of distilled  water  above
      the copper-cadmium granules to eliminate  entrapment of air.   Wash
      the column with 200 ml of dilute ammonium chloride solution.
      The column is then activated by passing through  the column
      100 ml of a solution composed of 25 ml of a 1.0-mg/Jl N03-N
      standard and 75 ml of ammonium chloride-EDTA solution.   Use a
      flow rate between 7 and 10 ml/min.
Ammonium chloride-EDTA solution:  dissolve 13 g ammonium chloride and

-------
Figure 3-22.    Nitrate reduction column
   10 cm
                 80-85 ml
            2 cm
   25 cm
1
18
\
5 cm
\
i
  3 cm I.D.
3.5 mm I.D.
  Cu / Cd
                           GLASS WOOL PLUG
                                  kCLAMP
                               TYGON TUBING
                 3-175

-------
      1.7 g disodium ethylenediamine tetracetate in 900 ml of distilled
      water.  Adjust the pH to 8.5 with cone,  ammonium hydroxide and
      dilute to 1 &.

Dilute ammonium chloride-EDTA solution:  dilute 300 ml of ammonium chlo-
      ride-EDTA solution to 500 ml with distilled water.

Color reagent:  dissolve 10 g sulfanilamide and 1 g N_ (.1-naphthyl)-
      ethylenediamine dihydrochloride in a mixture of 100 ml cone.
      phosphoric acid and 800 ml of distilled water and dilute to 1 
      with distilled water.

Zinc sulfate solution:  dissolve 100 g ZnS04    TH20 in distilled water
      and dilute to 1 .

Sodium hydroxide solution, 6 N_:  dissolve 2^0 g NaOH in 500 ml distilled
      water, cool, and dilute to 1 .

Ammonium hydroxide, cone.

Dilute hydrochloric acid, 6 N_:  dilute 50 ml of cone. HC1 to 100 ml with
      distilled water.

Copper sulfate solution, 2 percent:  dissolve 20 g of CuSOij   5^0 in
      500 ml of distilled water and dilute to 1 &.
Stock nitrate solution:  dissolve 7-218 g KNOa  in distilled water and
      dilute to 1000 ml.  Preserve with 2 ml of chloroform per liter.
      This solution is stable for at least 6 months.  1.0 ml = 1.00 mg
      NOa-N.
Standard nitrate solution:  dilute 10.0 ml of nitrate stock solution
      to 1000 ml with distilled water.  1.0 ml = 0.01 mg NOs-N.
Stock nitrite solution:  dissolve 6.072 g KNOa  in 500 ml of distilled
      water and dilute to 1000 ml.  Preserve with 2 ml of chloroform
      and keep under refrigeration.  This solution remains stable for
      approximately 3 months.  1.0 ml = 1.00 mg N02-W.
Standard nitrite solution:  dilute 10.0 ml of stock nitrite solution
      to 1000 ml with distilled water.  1.0 ml = 0.01 mg N02-N.
Using standard nitrate solution prepare the following standards in
      100-ml volumetric flasks:
      Cone., mg-N03-N/          ml of Standard Solution/100.0 ml

            0.00                               0.0
            0.05                               0.5
            0.10                               1.0
            0.20                               2.0
            0.50                               5.0
            1.00                              10.0
Procedure
            The presence of turbidity or suspended .solids can affect
the operation of the nitrate reduction column.   This material should


                               3-176

-------
be removed by one of the following methods:
      a_.  Filter sample through a glass fiber or a 0.^5-y membrane
          filter.
      b_.  Add 1 ml zinc sulfate solution to  100 ml of sample and mix
          thoroughly.  Add Q.k to 0.5 ml sodium hydroxide solution to
          obtain a pH of 10.5 as determined  with a pH meter.  Let the
          treated sample stand a few minutes to allow the heavy
          flocculent precipitate to settle.   Clarify by filtering
          through a glass fiber filter or a  0.1*5-y membrane filter.
            If oil or grease is known or thought to be present, it
should be removed by extraction.  Adjust the pH of 100-ml filtered
sample to 2 with cone.  HC1.   Extract the oil and grease with two 25-ml
portions of freon (or chloroform).
            Adjust the pH of the sample between 5 and 9 using either
cone. HC1 or cone. NH^OH.  To a 25-ml sample, or an aliquot diluted
to 25 ml, add 75 ml ammonium chloride-EDTA solution and mix.  The
sample pH should be 8.5.
            Pour the sample into the column  and adjust the flow to
7 to 10 ml per minute.   Discard the first 25 ml of sample and collect
the remainder of the sample in the original  sample flask.
NOTE:  The sample obtained from the column should not be held more
       than 15 min prior to color development to minimize nitrite
       oxidation.
            To 50 ml of reduced sample, add  2.0 ml color reagent.   The
color requires 10 min for development and is stable for 2 hr.   Measure
the sample absorbance at 5^0 nm relative to  a reagent blank.
NOTE:  If the sample concentration exceeds 1.0 mg N03-NA|, , the
       remainder of the reduced sample should be appropriately diluted.
       Develop the color as indicated above  and record the sample
       absorbance.
            Standards should be treated the  same as samples through-
out the entire procedure.  In addition, at least one nitrite standard
should be compared to a reduced nitrate standard at the same con-
centration to verify the efficiency of the reduction column.
Calculations
            Prepare a standard curve by plotting the absorbance of
standards against the nitrate and/or nitrite concentration.   Compute
sample concentration by comparing sample absorbance with the standard
                               3-177

-------
curve.
            If less than 25 ml of sample  is  used  for the  analysis, the
following equation should be used:
                   mg N02 + N03-N/ = -^	_
                                      ml  sample used
where
     A = concentration of nitrate from standard curve
                               3-178

-------
                                NITROGEN
                                (Nitrite)

                      Sample Collection and Storage

            Handling and storage requirements for  nitrite  samples  are
the same as nitrate samples.  Samples may  be collected and stored  in
either glass or plastic containers.   Environmental Protection Agency
manuals '7 suggest that sulfuric acid can  be used  as a preservative for
nitrite in water.  However, soils literature5 suggest that acid can
cause the conversion of nitrite.  It is recommended that water samples
be maintained at HC with minimum atmospheric contact.  Samples should
be processed as soon as possible.  The volume of sample required will
range from 20 to 100 ml depending on whether an automated  or  manual
method is used (.Figure 3-23).
            Nitrite may be determined using Methods 2,  3,  or  h in  the
Nitrate-Nitrogen section (Nitrite Methods  1, 2, and 3). The  only
alteration of these procedures is to omit  the use  of the reduction
column or the reducing agent and to use nitrite standards  rather than
nitrate standards.

              Procedures- for Water Samples (Wl, W2, S1A)
Method 1:  See Nitrate Method 2,  Colorimetric,  Automated,  Cadmium
           Reduction.
           Omit use of cadmium reduction column and proceed  as  indicated.
Method 2:  See Nitrate Method 3,  Colorimetric,  Automated,  Hydrazine
           Reduction.1
           Omit use of hydrazine  sulfate and proceed as  indicated.
Method 3:  See Nitrate Method k,  Colorimetric,  Manual, Cadmium
           Reduction.l
           Omit use of cadmium reduction column and proceed  as  indicated.
Method k:  Colorimetric, Manual1
            This procedure is a manual adaptation of Method  1.  Nitrite
is diazotized and coupled with napthyl-ethylenediamine dihydrochloride
                                3-179

-------
                Figure 3-23.     Handling and storage of samples for nitrite analysis

CORE SAMPLE
1

WATER SAMPLE JDREDGE SAMPLE CORE SECTION
1 * ,1
4 * * *
ACIDIFY FILTER N0 T STORE WET
1 1 	
STORE ACIDIFY |
1
STORE
1 '
*


14
ANALYZE ANALYZE ANALYZE ANALYZE
LO (Wl) (W2) (S1A) (SID)
H
O SAMPLE DESIGNATION
PURPOSE
CONTAINER
SAMPLE TREATMENT
PRESERVATIVE
STORAGE TIME
DIGESTION SOLUTION
Wl W2 W3 S1A SID
Total Water Soluble Used in Mobile Total
Cone. Water Elutriate Cone. Sediment
Cone. Cone.
G,P G.P G,P G,P G,P
None Filter None None None
HjSOi, H2SO None lteC ')C
pH<2 pH<2 (Minimize Air Contact. Keep Field Moist.)
li'C AC
W hr i8 hr <1 w 
-------
to produce a reddish-purple dye.  The sample absorbance is then measured
colorimetrically.

Apparatus
Spectrophotometer equipped with 1 cm or larger cells for use at 5^0 nm

Nessler tubes, 50 ml, or volumetric flasks, 50 ml

Reagents
Distilled water free of nitrite and nitrate is to be used in preparation
      of all reagents and standards.
Buffer-color reagent:  to 250 ml of distilled water, add 105 ml cone.
      hydrochloric acid, 5-0 g sulfanilamide, and 0.5 g N. (l-napthyl)
      ethylenediamine dihydrochloride.   Stir until dissolved.   Add  136 g
      of sodium acetate, CHsCOONa   3 HaO,  and again stir until
      dissolved.  Dilute to. 500 ml with distilled water.  This  solution
      is stable for several weeks if stored in the dark.
Nitrite stock solution:  1.0 ml = 0.10 mg N02-N.   Dissolve 0.^926 g of
      dried anhydrous sodium nitrite (2^ hr in desiccator)  in distilled
      water and dilute to 1000 ml.  Preserve with 2 ml chloroform per
      liter.
Nitrite standard solution:  1.0 ml = 0.001  mg N02-N.  Dilute 10.0 ml of
      the stock solution to 1000 ml.
Procedure
            If the sample has a pH greater  than 10 or a total alkalinity

greater than 600 mgA, adjust the sample pH to 6 with the addition  of
1:3 HC1.
            Filter the sample, if necessary, through a 0.^5-V pore-
size filter to remove suspended solids and  turbidity.
            Place 50 ml of sample, or an aliquot diluted to 50  ml,  in  a
50-ml Nessler tube.  Do not process samples further until the nitrite
standards are ready.
            Prepare a series of nitrite standards as suggested  below:
      ml of Standard Solution      Cone., When Diluted to 50 ml,
      1.0 ml = 0.001 mg N02-N      	mg/ of N02-N	
                0.0                           (Blank)
                0.5                            0.01
                1.0                            0.02
                1.5                            0.03
                2.0                            O.Oh
                3.0           '                 0.06
                ^.0                            0.08
                5.0                            0.10
               10.0                            0.20
                                3-l8l

-------
            Add 2 ml of buffer color reagent to each sample  and  standard.
Mix and allow 15 min for color development.   The resultant solution
should have a pH between 1.5 and 2.0.
            Measure the absorbance of the sample at  5^0  nm relative to
the blank.
Calculations
            Prepare a standard curve by plotting standard nitrite
concentration versus absorbance.  Compare the measured sample  absorbance
to the standard curve to determine the sample nitrite concentration.
            If a sample aliquot is diluted to 50 ml, calculate the
nitrite concentration as follows:
                                 from standard curve x 50
                       /? =
                2  '                ml sample used
                                3-182

-------
                  Procedure for Sediment Samples (.SID)

            This procedure for nitrate plus nitrite consists  of heating
the sediment slurry and then centrifuging out the solids.   The liquid
phase is then analyzed for nitrate plus nitrite.  This  separation pro-
cedure is "based on the high solubility of nitrates and  nitrites "but
should be considered operationally defined.  The apparatus  and reagents
vill depend upon which, of the methods listed in the nitrate and nitrite
sections for water analysis is used.
Procedure
            Weigh, a 0.5- to 1.0-g sample of the wet sediment.   Transfer
to a 200-ml Erlenmeyer flask and add  50 ml distilled water  and 3 to  ^
drops of cone, sulfuric acid.  This treatment will preserve the sample
for 2h hr if necessary.
            Add 50 ml distilled water to the acidified  slurry and boil
the sample for 15 min.  Since the procedure is operationally  defined,
the heating time should be standardized for all samples.
            Transfer the sample to a  centrifuge tube and  centrifuge
the slurry at 2000 rpm .for 5 to 10 min.  Decant the liquid  phase into
a 200-ml volumetric flask.
            Add 50 ml distilled water to the solids in  the  centrifuge
tube and thoroughly mix the sample.  Centrifuge for 5 to  10 min at
2000 rpm.  Decant the wash into the volumetric flask.
            Repeat the washing procedure a second time  and  add the
wash to the volumetric flask.  Dilute the sample to volume  with
distilled water.  Filter the sample through a O.U5-y pore-size
membrane filter.
            Analyze the sample using  one of the nitrate plus  nitrite
procedures in the water analysis section.  If it is necessary to know
the nitrate concentration, the sample should be analyzed  twice:  once
for nitrate plus nitrite using the cadmium reduction column or the
hydrazine reduction method, and a second time for nitrite by  omitting
the use of the cadmium reduction column or hydrazine sulfate.   Nitrate
can then be calculated by subtraction.

                                3-183

-------
            If the sample is to be analyzed for nitrate with the Brucine
method, the salt modification recommended for use with seawater samples
should be used to correct for the high salt concentration that may be
leached from the sediment samples.
Calculations
            Determine the nitrate plus nitrite concentration of the
sample leachate using the appropriate standard curve.   Calculate the
nitrate plus nitrite concentration of the sediment sample as follows:
         nitrate + nitrite mg N/kg (wet basis) =
where
     x = nitrate plus nitrite concentration in sample,
     y = sample volume, A (0.2 & as described)
     g = wet weight of sediment sample, g

         nitrate + nitrite mg N/kg (dry basis) =
where
     x = nitrate plus nitrite concentration in sample,
     y = sample volume, & (0.2  as described)
     g = wet weight of sediment sample,  g
    %S = percent solids in sediment (as  decimal fraction)
                                3-181*

-------
                               NITROGEN
                           (Total Kjeldahl)

                     Sample Collection and Storage

            Total Kjeldahl nitrogen (TKN) samples may be collected and
held in either glass or plastic containers.  Samples may be preserved
with sulfuric acid (pH < 2) at ^C and should be analyzed within 2.h hr.
Longer storage times have been reported at high TKN concentrations.  A
flowchart for sample handling is presented in Figure 3-2.14.

              Procedures for Water Samples (,W1, W2, S1A)

                                                          189
Method 1:  Colorimetric, Semiautomated with Block Digester ' '
            The procedure consists of two parts.  The sample is initially
digested with a sulfuric acid-potassium sulfate-mercury sulfate solution
to convert organic nitrogen to ammonia.  The sample is then analyzed for
total ammonia (.original ammonia plus covered organic nitrogen).  The
applicable range is 0.1 to 20 mg TKN-N/&.
Apparatus-
Block digestor-HO and digestion tubes
Technicon manifold for ammonia (Figure 3-25)
Chemware TFE (.Teflon boiling stones), Markson Science, Inc., Box 767,
      Delmar, California 9201^
Reagents
Mercuric sulfate:  dissolve 8 g red mercuric oxide, HgO, in 50 ml of
      I:h sulfuric acid (.10 ml cone. HzSOitiUo ml distilled water) and
      dilute to 100 ml with distilled water.
Digestion solution (.sulfuric acid-mercuric sulfate-potassium sulfate
      solution):  dissolve 133 g of KaSOi, in 700 ml of distilled water
      and 200 ml of cone. J^SOi*.  Add 25 ml of mercuric sulfate solution
      and dilute to 1 .
Sulfuric acid solution (.U percent):  add Uo ml of cone, sulfuric acid
      to 800 ml of ammonia-free distilled water, cool, and dilute to
      1 .
Stock sodium hydroxide C20 percent):  dissolve 200 g of sodium hydroxide
      in 900 ml of ammonia-free distilled water and dilute to 1 .
                                3-185

-------
                   Figure 3-2U.    Handling  and storage of samples for  TKN analysis

^^"""^""
ACIDIFY
T r
STORE
1
DIGEST
1
ANALYZE


CORE SAMPLE
4
i^^___  ^
WATER SAMPLE DREDGE SAMPLE CORE SECTION
4 1 *
* t
FILTER N0 T^3JMENT STORE WET
1
ACIDIFY
1
STORE
i '
^ ^


1 1 I
ANALYZE ANALYZE ANALYZE
(W2) (S1A) (SID)

SAMPLE DESIGNATION
PURPOSE
CONTAINER
SAMPLE TREATMENT
PRESERVATIVE
STORAGE TIME
DIGESTION SOLUTION
Wl U2 W3 S1A SID
Total Water Soluble Used in Mobile Total
Cone. Water Elutriate Cone. Sediment
Cone. Cone.
G,P G,P G,P G,P G,P
None Filter None None None
H2SOi, HzSOt, None 
-------
                 Figure  3-25.    AAII  cartridge  for ammonia determinations with TKN digests
OJ

H
oo
                      37C


          5 Turns  157-B273-03

                     OOOQ
  TO PUMP
                           660 nm
10 Turns
00000
157-
20 T
000(
10 Turns
J
> ;
 -

^r

-^f
~~
116-0489-01
  -
B089
urns
)OOO







-^


METER
m ^ 	
F/C x 1 . 5 mm 1 D
PROPORTION IN
PUMP
Gry Gry
Blk Blk
Red Red
Orn Yel
Blk Blk
Red Red
Orn Yftl
Blk Blk
Orn YeJ
Gry Gry
	 . 	
G
ml /mi n .
1 .0 k% H2SOi,
0.32 AIR
0.80 DILUENT WATER
*SAMPLE ^
116-
0.32 AIR
0.80 WORKING BUFFER
ARESAMPLE
fTO PH
SAMP
PT2


BOOO
\
0.32 SALICYLATE-NITROPRUSSIOE
0.16 HYPOCHLORITE
1.0 WASTE



OSPHORUS
LE LINE
'
                                                                          See Table  3'l8 for range selection.

-------
Stock sodium potassium tartrate solution (20 percent):   dissolve 200 g
      potassium tartrate in about 800 ml of ammonia-free distilled
      water and dilute to 1 fc.
Stock buffer solution:  dissolve 13^.0 g of sodium phosphate, dibasic,
      NaaHPOij, in about 800 ml of ammonia-free water.   Add 20 g of
      sodium hydroxide and dilute to 1 .
Working buffer solution:  combine the reagents in the  stated order;
      add 250 ml of stock sodium potassium tartrate solution to 200  ml
      of stock buffer solution and mix.  Add xx ml sodium hydroxide
      solution and dilute to 1 H.  The exact volume of sodium hydroxide
      solution (xx ml) will vary with the expected nitrogen concentration
      as- indicated in the last column in Table 3-18-
Sodium salicylate/sodium nitroprusside solution:  dissolve 150 g of  so-
      dium salicylate and 0.3 g of sodium nitroprusside in about 600 ml
      of ammonia-free water and dilute to 1 &.
Sodium hypochlorite solution:  dilute 6.0 ml sodium hypochlorite
      solution (.Clorox) to 100 ml with ammonia-free distilled water.
Ammonium chloride, stock solution:  dissolve 3.819 g NHi,Cl in distilled
      water and bring to volume in a l- volumetric flask.   1 ml =
      1.0 mg NH3-N.
Procedure
            Transfer 25 ml of sample to a digestion tube and add 5 ml
digestion solution.  Mix sample with a vortex mixer to avoid super-
heating during digestion.  Add k to 5 Teflon boiling stones.
            With block digester in manual mode,  set low and high
temperature at l6oC and preheat unit to l6oC.   Place tubes in digestor
and switch to automatic mode.  Set low temperature timer for 1 hr.
Reset high temperature to 380C and set timer for 2-1/2 hr.
            Allow ingested samples to cool to room temperature and
dilute to 25 ml with ammonia-free water.
            Prepare an ammonia manifold as indicated in Figure 3-2$.
Check all reagent containers to ensure an adequate supply.
            Excluding the salicylate line, place all reagent lines in
their respective containers, connect the sample  probe  to the sampler,
and start the proportioning pump.
            Flush the sampler wash receptacle with approximately
25 ml U.O percent sulfuric acid.
            When reagents have been pumping for  at least 5 min, place
the salicylate line in its respective container  and allow the system to
                                3-188

-------
                                                    Table 3-l8

                              Operating Characteristics for TKN AutoAnalyzer Manifold

                                               CONCENTRATION RANGES

                                                     (NITROGEN)

No.
1
2
3
k
5
6
Initial
Sample Line
.80 (RED/RED)
.80 (RED/RED)
.16 (ORN/YEL)
.16 (ORN/YEL)
.16 (ORN/YEL)
.16 (ORN/YEL)
Diluti
Sample
Diluent Line
.80 (RED/RED)
.80 (RED/RED)
.80 (RED/RED)
.80 (RED/RED)
.80 (RED/RED)
.80 (RED/RED)
on Loops
Res
Resample Line
.32 (BLK/BLK)
.32 (BLK/BLK)
.32 (BLK/BLK)
.32 (BLK/BLK)
.16 (ORN/YEL)
.16 (ORN/YEL)
ample
Di luent Line
.80 (RED/RED)
.80 (RED/RED)
.80 (RED/RED)
.80 (RED/RED)
.80 (RED/RED)
.80 (RED/RED)
Approx.
std. cal.
setting
700
100
700
100
700
100
Range
PPM N
(10%)
0-0.5
0-1.5
0-1
0-5
0-2
0-10
ml stock NaOH
per 1 i ter
working buffer
sol ution
250
250
120
120
80
80
H
CO
MD

-------
equilibrate.   If a precipitate forms after the addition of salicylate,
the pH is too low.  Immediately stop the proportioning pump and flush
the coils with water using a syringe.   Before restarting the system,
check the concentration of the sulfuric acid solutions and/or the
working buffer solution.
            To prevent precipitation of sodium salicylate in the waste
tray, which can clog the tray outlet,  keep the nitrogen flow cell pump
and the nitrogen colorimeter "To Waste" tube separate from other lines
or keep tap water flowing in the waste tray.
            Place ammonia standards in sample tray in order of
decreasing concentration.  Complete loading of sample tray with
digested samples.  When a stable baseline has been obtained, switch
sample line to sampler and start analysis.  Use a itO/hr, k:l cam for
the Mil.
Calculations
            Prepare standard curve by plotting peak heights of
processed standards against concentration values.   Compute concentra-
tions by comparing sample peak heights with standard curve.

Method 2:  Manual Colorimetric, Titrimetric
            The first step in this method consists of digestion and
distillation.  Ammonia, both original ammonia and organic nitrogen
converted to ammonia, is distilled into a boric acid solution.   The
ammonia can then be quantitated by nesslerization or sulfuric acid
titration.  The nesslerization procedure is suitable to ammonia con-
centrations below 1 mg NHa-K/X- and the titrimetric procedure is
suitable to ammonia concentrations above 1 mg NHs-N/A.
Apparatus
Digestion apparatus:  a Kjeldahl digestion apparatus with 800- or 1000-
      ml flasks and suction takeoff to remove SO3 fumes and water
Distillation apparatus:  the macro Kjeldahl flask is connected to a
      condenser and an adaptor so that the distillate can be collected.
      Micro Kjeldahl steam distillation apparatus is commercially
      available.
Spectrophotometer for use at HOO to U25 nm with a light path of 1 cm  or
      longer
                                 3-190

-------
Reagents

Distilled water should be free of ammonia.   Such water is best prepared
      by the passage of distilled water through an ion exchange column
      containing a strongly acidic cation exchange resin mixed with a
      strongly basic anion exchange resin.   Regeneration of the column
      should be carried out according to the manufacturer's instructions.

NOTE:  All solutions must be made with ammonia-free water.

Mercuric sulfate solution:  dissolve 8 g red mercuric oxide, HgO,  in
      50 ml of l:h sulfuric acid (10.0 ml cone. Ha SOi* :kO ml distilled
      water) and dilute to 100 ml with distilled water.

Sulfuric acid-mercuric sulfate-potassium sulfate solution:  dissolve
      267 g K2S(\ in iSOO ml distilled water and HOO ml cone.  feSOn.
      Add 50 ml mercuric sulfate solution and dilute to 2 & with
      distilled water.

Sodium hydroxide-sodium thiosulfate solution:  dissolve 500 g NaOH and
      25 g Na2S203  5H20 in distilled water and dilute to 1 .

Mixed indicator:  mix 2 volumes of 0.2 percent methyl red in 95 percent
      ethanol with 1 volume of 0.2 percent methylene blue in ethanol.
      Prepare fresh every 30 days.
Boric acid solution:  dissolve 20 g boric acid, H3B03, in water and
      dilute to 1  with distilled water.
Sulfuric acid, standard solution:  (0.02 N_) 1 ml = 0.28 mg NH3-N.
      Prepare a stock, solution of approximately 0.1 N_ acid by diluting
      3 ml of cone. H2S0lt (sp. gr. I.Qk] to 1  with C02-free distilled
      water.  Dilute 200 ml of this solution to 1 Jl with C02-free
      distilled water.  Standardize the approximately 0.02 N_ acid  so
      prepared against 0.0200 N_ Na2C03 solution.  This last solution
      is prepared by dissolving 1.060 g anhydrous Na2C03, oven dried
      at 1^0C, and diluted to 1  with C02-free distilled water.

NOTE:  An alternate and perhaps preferable method is to standardize
       the approximately 0.1 N_ H2S04 solution against a 0.100 IT
       Na2C03 solution.  By proper dilution, the 0.02 N_ acid can
       then be prepared.

Ammonium chloride, stock solution:  1.0 ml = 1.0 mg NH3-N.  Dissolve
      3.819 g NHijCl in water and make up to 1  in a volumetric flask
      with distilled water.

Ammonium chloride, standard solution:  1.0 ml = 0.01 mg NH3-N.
      Dilute 10.0 ml of the stock solution with distilled water to 1 
      in a volumetric flask.
Nessler reagent:  dissolve 100 g of mercuric iodide and 70 g potassium
      iodide in a small volume of distilled water.  Add this mixture
      slowly, with stirring, to a cool solution of 160 g of NaOH in
      500 ml of distilled water.  Dilute the mixture to 1 .  The
      solution is stable for at least 1 year if stored in a pyrex
      bottle out of direct sunlight.

                                 3-191

-------
Procedure
            The distillation apparatus  should be presteamed before use by
distilling a 1:1 mixture of distilled water  and sodium hydroxide-sodium
thiosulfate solution until the distillate is ammonia  free.  This opera-
tion should be repeated each time the apparatus is  out of  service long
enough to accumulate ammonia (usually k hr or more).
            Macro Kjeldahl digestion.   Place a measured  sample  or the
residue from the distillation in the ammonia determination (for organic
Kjeldahl only) into an 800-ml Kjeldahl  flask. The  sample  size  can be
determined from the following table:
      Kjeldahl Nitrogen in Sample                    Sample Size
      	mg/jj.	                       ml
                0-5                                 500
                5-10                                 250
               10 -  20                                 100
               20 -  50                                 50.0
               50 -  500                                25.0
            Dilute the sample, if required,  to 500  ml with distilled
water and add 100 ml sulfuric acid-mercuric  sulfate-potassium sulfate
solution.  Evaporate the mixture in the Kjeldahl apparatus until SOs
fumes are given off and the solution turns colorless  or  pale yellow.
Continue heating for 30 additional minutes.   Cool the residue and add
300ml distilled water.
            Make the digestate alkaline by careful  addition of  100 ml
of sodium hydroxide-thiosulfate solution without mixing.
NOTE:  Slow addition of the heavy caustic solution  down  the tilted
       neck of the digestion flask will cause heavier solution  to
       underlay the aqueous sulfuric acid solution  without loss of
       free ammonia.  Do not mix until  the digestion  flask has  been
       connected to the distillation apparatus.
            Connect the Kjeldahl flask  to the condenser with the tip
of the condenser or an extension of the condenser tip below the level
of the boric acid solution in the receiving  flask.
            Distill 30 ml at the rate of 6 to 10 ml/min  into  50 ml of
2 percent boric acid contained in a 500-ml Erlenmeyer flask.
            Dilute the distillate to 500 ml  in the  flask.  These
flasks should be marked at the 350- and the  500-ml  volumes.   With  such
                                 3-192

-------
marking, it is not necessary to transfer the distillate to volumetric
flasks.  For concentrations above 1 mg/, the ammonia can be  determined
titrimetrically.  For concentrations below this value, it is  determined
color imetrically.
            Micro Kjeldahl digestion.   Place 50.0 ml  of sample or  an
aliquot diluted to 50 ml in a 100-ml Kjeldahl flask and add 10 ml
sulfuric acid-mercuric sulfate-potassium sulfate solution.
            Evaporate the mixture in the Kjeldahl apparatus until  S03
fumes are given off and the solution turns colorless  or pale  yellow.
Then digest for an additional 30 min.   Cool the residue and add 30 ml
distilled water.
            Make the digestate alkaline by careful addition of 10  ml of
sodium hydroxide-thiosulfate solution without mixing.   Do not mix  until
the digestion flask has been connected to the distillation apparatus.
            Connect the Kjeldahl flask to the condenser with  the tip
of condenser or an extension of the condenser tip below the level  of
the boric acid solution in the receiving flask or 50-ml short-form
Nessler tube.
            Steam distill 30 ml at the rate of 6 to 10 ml/min into
5 ml of 2 percent boric acid.
            Dilute the distillate to 50 ml.  For concentrations above
1 mg/, the ammonia can be determined titrimetrically.   For concentra-
tions below this value, it is determined colorimetrically.
            a_.  Titrimetric determination:   add 3 drops of the mixed
                indicator to the distillate and titrate the ammonia
                with the 0.02 N_ H2SOi,, matching the endpoint  against
                a blank containing the same volume of distilled water
                and H3B03 solution.
            b_.  Colorimetric determination:  prepare  a series of
                Nessler tube standards as follows:
                          ml of Standard
                      1.0 ml = 0.01 mg NH^-N    mg NHa-N/50.0 ml
                                0.0                   0.0
                                0.5                   0.005
                                1.0                   0.010
                                2.0                   0.020
                                ^.0                   O.OUO
                                5.0                   0.050
                                8.0                   0.080
                               10.0                   0.10
                                 3-193

-------
                Dilute each tube to 50 ml with ammonia-free water,  add
                1 ml of Nessler reagent,  and mix.   After 20 min,  read
                the absorbance at 1^25 nm against the blank.  From the
                values obtained for the standards,  plot absorbance  vs.
                mg NHs-N for the standard curve. Develop color in  the
                50-ml diluted distillate in exactly the same manner
                and read mg NHs-N from the standard curve.
            It is not imperative that all standards be treated in the
same manner as the samples.  It is recommended that at least two
standards (high and low) be digested, distilled, and compared to  similar
values on the curve to ensure that the digestion-distillation technique
is reliable.  If treated standards do not agree with untreated standards,
the operator should find the cause of the apparent  error before pro-
ceeding.
Calculations
            If the titrimetric procedure is used, calculate total
Kjeldahl nitrogen, in mg/&, in the original sample  as follows:
                    T0> ^ = (A - B)N x F x 3.QOO
                                         Q
where
     A = volume of standard 0.020 W_ H SO  solution  used in titrating
         sample, ml
     B = volume of standard 0.020 N_ H2SOi, solution  used in titrating
         blank, ml
     N = normality of sulfuric acid solution
     F = milliequivalent weight of nitrogen (lU mg)
     S = volume of sample digested, ml
            If the Nessler procedure is used, calculate the total
Kjeldahl nitrogen, in mg/, in the original sample  as follows:
                       miri.T    ,.   A X 1000   B
                       TKW, mg/ =	 x -
where
     A = NHa-N read from curve, mg
     B = total distillate collected including the HijBOs, ml
     C = distillate taken for nesslerization, ml
     D = original sample size, ml
                               3-19**

-------
            Calculate organic Kjeldahl nitrogen, in mg/Jl, as follows:
               Organic Kjeldahl Nitrogen = TKN - (NH3-N)

Method 3:  Colorimetric, Automated Phenate
            This procedure is a completely automated method to determine
total Kjeldahl nitrogen in the range of 0.05 to 2.0 mg TKN-N/fc.   The
sample is digested with a sulfuric acid-potassium sulfate-mercuric
sulfate solution.  The digestate is then treated successively with
alkaline phenol, sodium hypochlorite, and sodium nitroprusside.   Sample
absorbance is then measured to quantitate the total Kjeldahl nitrogen
concentration.
            The user is cautioned that the manifolds for this procedure
are rather complex.  If this method is selected, the special cautions
provided in the procedure section should "be closely adhered to.
Apparatus
Technicon autoanalyzer consisting of:
      a_.  Sampler II, equipped with continuous mixer
      b_.  Two proportioning pumps
      c_.  Manifold I
      cL  Manifold II
      e_.  Continous digester
      _f.  Planatary pump
      .  5-gal carboy fume-trap
      h.  80C heating bath
      i^.  Colorimeter equipped with 50-mm tubular flow cell and 630-nm
          filters
      j_.  Recorder equipped with range expander
      k_.  Vacuum pump
Reagents
Distilled water:  special precaution must be taken to ensure that
      distilled water is free of ammonia.  Such water is prepared
      by passage of distilled water through an ion exchange column
      comprised of a mixture of both strongly acidic cation and strongly
      basic anion exchange resins.  Furthermore, since organic contami-
      nation may interfere with this analysis, use of the resin Dowex
      XE-T5 or equivalent which also tends to remove organic impurities
      is advised.  The regeneration of the ion exchange column should

                                3-195

-------
      be carried out according to the instruction of the manufacturer.

NOTE:  All solutions must be made using ammonia-free water.

Sulfuric acid:  as it readily absorbs ammonia, special precaution must
      also be taken with respect to its use.   Do not store bottles
      reserved for this determination in areas of potenial ammonia
      contamination.

EDTA (.2 percent solution):  dissolve 20 g disodium ethylenediamine
      tetraacetate in 1  of distilled water.   Adjust pH to  10.5  - 11
      with NaOH.

Sodium hydroxide (30 percent solution):  dissolve 300 g NaOH in 1 A of
      distilled water.

NOTE:  The 30 percent sodium hydroxide should  be sufficient  to neutralize
       the digestate.  In rare cases it may be necessary to  increase the
       concentration of sodium hydroxide in this solution to ensure
       neutralization of the digested sample in the manifold at the
       water-jacketed mixing coil.

Sodium nitroprusside (0.05 percent solution):   dissolve 0.5  g Na2Fe(CN)s
      NO    2H20 in 1 t, distilled water.

Alkaline phenol reagent:  pour 550 ml liquid phenol (.88 to 90 percent)
      slowly with mixing into 1  of 1*0 percent (>00 g/&) NaOH.  Cool
      and dilute to 2 I with distilled water.

Sodium hypochlorite (l percent solution):   dilute commercial Clorox,
      200 ml to 1  with distilled water.   Available chlorine level
      should be approximately 1 percent.  Due  to the instability  of this
      product, storage over an extended period should be avoided.

Digestant mixture:  place 2 g red HgO in a 2- container. Slowly add,
      with stirring, 300 ml of acid water (100 ml ^SQ^ + 200 ml  H20)
      and stir until cool.  Add 100 ml 10 percent (.10 g per  100 ml)
      K2S04.   Dilute to 2  with cone, sulfuric acid (approximately
      500 ml  at a time, allowing time for cooling).  Allow U hr for
      the precipitate to settle or filter through glass fiber filter.

Stock solutions:  dissolve k.l6l9 g of predried (l hr at 105C) ammonium
      sulfate in distilled water and dilute to 1.0  in a volumetric
      flask.   1.0 ml = 1.0 mg N.
Standard solution:  dilute 10.0 ml of stock solution to 1000 ml.
      1.0 ml = 0.01 mg N.  Using the standard  solution, prepare the
      following standards in 100-ml volumetric flasks:

      ml Standard Solution/100 ml                   Cone., mg N/
                  0.0                                   0.00
                  0.5                                   0.05
                  1.0                                   0.10
                  2.0                                   0.20
                  U.O                                   0.1+0
                  6.0                                   0.60
                  8.0                                   0.80
                 10.0                                   1.00
                 15.0                                   1.50
                 20.0                                   2.00
                                 3-196

-------
Procedure
            Set up the Technicon manifolds  as  shown  in Figures  3-26
through 3-28.
NOTE:  In the operation of manifold No.  1,  the control of  four  key
       factors is required to enable manifold  No.  2  to receive  the
       mandatory representative feed.  First,  the  digestant flowing
       into the pulse chamber (PCl) must be bubble free; otherwise, air
       will accumulate in A-7 thus altering the ratio of  sample to
       digestant in digestor.  Second, in maintaining even flow from
       the digestor helix, the peristaltic  pump must be adjusted to cope
       with differences in density of the digestate  and the wash water.
       Third, the sample pickup rate from the  helix  must be precisely
       adjusted to ensure that the entire sample is  aspirated into the
       mixing chamber.  Finally, the contents  of the mixing chamber
       must be kept homogeneous by the proper  adjustment of the air
       bubbling rate.
NOTE:  In the operation of manifold No.  2,  it  is important in the
       neutralization of the digested sample to adjust the concentration
       of the NaOH so that the waste from the  C-3  debubbler is  slightly
       acidic to Hydrion B paper.
NOTE:  The digestor temperature is 390C for the first stage and 36oC
       for the second and third stages.
            Allow both colorimeter and recorder to warm up for  30 min.
Run a baseline with all reagents, feeding distilled  water  through the
sample line.  Adjust dark current and operative opening on colorimeter
to obtain stable baseline.
            Set sampling rate of Sampler II at 20  samples  per hour,
using a sample to wash ratio of 1 to 2  (l-min  sample, 2-min wash).
            Arrange various standards in sampler cups in order  of
increasing concentration.  Complete loading of sampler tray with unknown
samples.  Switch sample line from distilled water  to sampler and begin
analysis.
Calculations
            Prepare standard curve by plotting peak  heights of  processed
standards against concentration values.   Compute concentration  of samples
by comparing sample peak heights with standard curve.
            It is suggested that any sample with a calculated concentra-
tion less than 10 percent of the sample  analyzed just prior to  it should
be rerun.
                                  3-197

-------
                         Figure 3-26.     AAI manifold 1 for the determination of TKN
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1 .60
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AIR* P O
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        PLANATARY
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                               DIGESTOR
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                                                                                                >For Sal t Water
                                                                                                   Samples ***
PROPORTIONING
    PUMP
                        * Air is scrubbed through 5N H2SOi

                       ** Teflon tubing or glass.

                      *** For fresh water samples use:

                             P    BI 2.90  WASH WATER
                                  PJ2.50  SAMPLE  	
                                 "G~l 2.00  DISTILLED H20
                                     1.20  AIR*
         o o

-------
                          Figure 3-27.   AAI manifold 2 for the  determination of TKW
MD
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0.80 DISTILLED WATER
1.20 AIR
2.90 SAMPLE _
1.20 EDTA
0.80 AIR
0.80 niSTIM pn UATFR
1.20 ALK. PHENOL
1.20 NaOCl
1.20 NITROPRUSSIDE
3-^0 WASTE 	
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RECORDER

-------
        Figure 3-28.     Continuous digester  for the automated  determination of total Kjeldahl nitrogen
    TO ASPIRATOR
ro
o
o
                                  CONTINUOUS DIGESTOR 5 MIXING CHAMBER ASSEMBLY
                          DILUTION WATER'"
                     ADJUSTABLE FLOW
                     'RENN1 PLANATARY
                          PUMP
       5  GALLON
       FUME CARBON TRAP
       HALF FILLED WITH
       1*0% NaOH
       CONNECTED TO
       ASPIRATOR
                        TO SUMPj
                           TO ASPIRATOR u
LINE DESIGNATION:

    Oxidized Sample

     ir Agitation

    Mixing Chamber Overflow

    Waste
    Feed to Manifold #2
                                              TO MANIFOLD #2

-------
                    Procedures for Sediment Samples

Method 1:  Kjeldahl Digestion
            This procedure is essentially the same as that used for TKN
analysis of water samples.  The sample is digested with a sulfuric acid-
potassium sulfate-mercury sulfate solution to convert organic nitrogen to
ammonia.  The digestate is made alkaline and the ammonia is distilled into
boric acid.  This solution is then analyzed for ammonia.
Apparatus
Digestion apparatus
Distillation apparatus
Reagents
Digestion solution:
      a_.  Dissolve 8 g red mercuric oxide, HgO, in 50 ml 1:5 HaSCK and
          dilute to 100 ml with distilled water.
      b_.  Prepare a second solution by dissolving 267 g KaSOi* in 1300 ml
          distilled water and 1*00 ml cone, sulfuric acid.  Add 50 ml of
          the mercuric sulfate solution and dilute to 2 &.
Phenolphthalein indicator solution:  either the aqueous (a) or alcoholic
      (t>) solution may be used:
      a_.  Dissolve 5 g phenolphthalein disodium salt in distilled water
          and dilute to 1 .  If necessary, add 0.02 N NaOH dropwise until
          a faint pink color appears.
      b_.  Dissolve 5 g phenolphthalein in 500 ml 95 percent ethyl alcohol
          or i&opropyl alcohol and add 500 ml distilled water.  Add 0.02
          N_ NaOH dropwise until a faint pink color appears.
Sodium hydroxide-sodium thiosulfate solution:  dissolve 500 g NaOH and
      25 g Na2S203   5H20 in distilled water and dilute to 1 .
Boric acid solution:   dissolve 20 g anhydrous boric acid, H3B03, in
      ammonia-free water and dilute to 1 .
Procedure
            Weigh out a 0.5- to 1.0-g aliquot of wet sediment.  Transfer
to a 800-ml digestion tube and add 100 ml digestion solution.  Boil until
white fumes appear.  Continue heating for an additional 30 min.  During
this time, the sample flask should be rotated occasionally.
            Cool sample to room temperature.  Add 500 ml ammonia-free
water and 0.3 ml phenolphthalein indicator.  Carefully add 25 to 30 ml
                                  3-201

-------
NaOH-thiosulfate solution to the Kjeldahl flask without mixing the
sample.  Place the flask on a digestion rack and mix.
NOTE:  If the solution is not red at this point, additional NaOH-
       thiosulfate solution must be added until a red  color appears.
            Distill over approximately 300 ml of liquid at a rate of  6
to 10 ml/min.  Collect the distillate in 50 ml 2 percent boric acid.
Dilute to 500 ml.
            Analyze the sample using one of the automated methods,
direct nesslerizatlon, or titration procedures for ammonia as described
in the water section.  Nesslerization should only be used when the
ammonia concentration is less than 1 mg/& and the titration procedure
should only be used when the ammonia concentration is  greater than
1 mg/.
Calculations
       Total Kjeldahl Nitrogen mg/kg (wet basis) = (xHy)(.1000)

       Total Kjeldahl Nitrogen mg/kg (dry basis) =
                                                     (
where
       x = ammonia concentration in distillate, mg/&
       y = total volume of distillate, & (0.5 as described)
       g = wet weight of sediment used, g
     % S = percent solids in sediment (as decimal fraction)

Method 2:  Block Digestion
            The sample is digested with a sulfuric acid-potassium
sulfate-mercury sulfate solution on a Technicon digestion block.   The
digest is diluted to volume and analyzed with an autoanalyzer.
Apparatus
Technicon BD-^0 block digestor
Technicon #11^-0009-02 test tube rack
Pyrex digestion tubes, 1 by 8 in.
Technicon autoanalyzer:
      a_.  Sampler with 30/hr 2:1 cam
      b_.  Proportionating pumps
                                 3-202

-------
      c_.  Colorimeter

      d_.  Recorder

      e_.  Digital printer (optional)

      _f.  Ammonia manifold as shown in Figures 3-13 and 3-1^
Teflon "boiling stones
Reagents
Ammonia-free water.

Digesting solution:  dissolve 2.Q g of HgO in 25 ml of 6 N H2SOt.   Then
      carefully add 200 ml of cone. HzSOt to 500 ml of water.   While the
      strong acid solution is still hot, dissolve 13^ g of K2S04 in it.
      Add the HgO solution.  Cool the solution, dilute to 1  ,  and store
      above 20C.  It is extremely important that precipitation of the
      K2SOi be avoided, as this will result in low recoveries  for  TKW.
      The digestion tube dilution water should be nitrogen and phosphorus
      free.

Reagents for automated dilution manifold (Figure 3-13) :   prepare the
      sampler wash solution by adding 35 ml of cone.  H2SOn to  500  ml of
      water and diluting to 1 9>.  Prepare the dilution manifold solution
      by diluting 12.5 ml of 10 N NaOH to 1 & with water.
Reagents for automated ammonia manifold (Figure 3-lU):  Prepare complex-
      ing reagent by dissolving 33 g of potassium sodium tartrate  and
      2k g of sodium citrate in 900 ml of water, diluting to 1 &,  and
      adding 0.25 ml of Brij-35 wetting agent (Technicon No. T21-0110).
Prepare alkaline phenol solution:  dissolve 83 g of phenol and  36  g of
      sodium hydroxide in 900 ml of water, cooling, and diluting to 1 &.
      Store the solution at kC.
Prepare sodium hypochlorite solution:  dilute 200 ml of Clorox  (5-25
      percent available 012) to 1  with water.
Prepare sodium nitroprusside reagent:  dissolve 0.5 g of sodium nitro-
      prusside in 900 ml of water and dilute to 1 JL   Store the solution
      at UC.
Standards:  Prepare stock nitrogen standard containing 0.100 mg N/ml
      by dissolving 1.050 g of glutamic acid, dried at 105C for 1 hr,
      in 900 ml of water.  Add 2 ml of cone. foSOi* and dilute  to 1  .
Procedure
            Weigh out 0.5 to 1.0 g of dry weight equivalent sediment and
transfer to a digestion tube.   Add 10 ml digestion solution and 2 to 3
Teflon boiling stones.  Mix samples well using a genie vortex mixer.
            Place samples in holder and place holder on the digestion
block.  Heat the samples for 1 hr at 200C and 1 hr at 370C (total
digestion time is 2 hr).


                                 3-203

-------
            Cool samples to room temperature and  dilute  to  75 ml with
distilled water.  Allow solids to settle and decant  sufficient  liquid
to fill autoanalyzer sample cups.
            Place standards in sample tray in order  of decreasing
concentration.  Continue filling tray with unknown samples.  After the
instrument has warmed up for 30 min and a stable  baseline has been
achieved, begin processing samples at the rate of 30/hr.
Calculations
            Determine the ammonia concentrations  in  the  digestates by
comparing sample peak height with the standard curve based  on the
instrument response to the standard ammonia solutions.   Calculate
the total Kjeldahl nitrogen concentration in the  same as follows:
                 TKN mg/kg (wet weight)  = Cx)(y)(lOOO)

                 TKN mg/kg (dry weight)  =
where
       x = ammonia concentration in digestate, mg/&
       y = volume of digestate, H (0.075 & as written)
       g = wet weight of sample, g
     % S= percent solids in sediment (as decimal fraction)
                                 3-20U

-------
                               NITROGEN
                               (Organic)

            A specific procedure is not provided for this parameter
                                         1  3
since it can be calculated mathematically '   or defined by sample
treatment.1'3  Mathematically, organic nitrogen can be calculated as
total Kjeldahl nitrogen minus the ammonia concentration of the sample.
Analytically, the sample is pretreated by distilling off the ammonia
at a pH of 9-5-  The residual is subjected to one of the total Kjeldahl
nitrogen methods described earlier and the result is termed organic
nitrogen.
                      Sample Handling and Storage

            Since organic nitrogen is defined as the difference betveen
total Kjeldahl nitrogen and ammonia nitrogen, samples should be handled
as discussed earlier for these parameters.   This information is presented
in Figure 3-12 for ammonia and Figure 3-2k for total Kjeldahl nitrogen.
Any of the water fractions, 1, 2, or S1A,  may be analyzed for organic
nitrogen.  However, it is recommended that nitrogen analyses be run on
wet sediment samples only.  Samples stored either dried or frozen may
be altered by the oxidation of nitrites and the absorption or volatili-
zation of ammonia.  In addition, the microbial population may alter the
organic nitrogen concentration.  To minimize these potential effects,
wet sediment samples should be used and processed as soon as possible.
This suggestion applies to all forms of nitrogen in sediments as indi-
cated in Figures 3-12, 3-l6, 3-23, and 3-2li.

                                Calculations
            Organic nitrogen is the difference between total Kjeldahl
nitrogen and ammonia nitrogen.
            Water samples:
           Organic nitrogen-N mg/ = TKN-N mg/& - NH3-N mg/
            Sediment samples:
         Organic nitrogen-N mg/kg = TKN-N mg/kg - NH3-N mg/kg

                                3-205

-------
                               References
1.  Environmental Protection Agency.   "Methods for Chemical Analysis  of
    Water and Wastes."  Environmental Monitoring and Support Laboratory,
    EPA; Cincinnati, Ohio (197*0.

2.  American Public Health Association.   Standard Methods for the
    Examination of Water and Wastewater  Including Bottom Sediments  and
    Sludges.   l*rth Edition.   APHA;  New York,  New York.   1193 p.  (1976).

3.  Environment Canada.   "Analytical Methods  Manual."  Inland Waters
    Directorate, Water Quality Branch; Ottawa, Canada (197*0-

k.  American Society for Testing Materials.   "Annual Book of ASTM
    Standards, Part 31,  Water."  American Society for Testing Materials;
    Philadelphia, Pennsylvania (1976).

5-  Hesse, P. R.  A Textbook of Soil Chemical Analysis.   Chemical
    Publishing Company;  New York,  New York.   520 p. (1971).

6.  Carter, M. J.  "Wastewater Sample Preservation Data for Twenty  Common
    Water Quality Parameters."  National Environmental Enforcement  Center;
    Denver, Colorado (1978).

7-  Environmental Protection Agency.   "Methods for Chemical Analysis  of
    Water and Wastes."  Environmental Monitoring and Support Laboratory,
    EPA; Cincinnati, Ohio (1979).
8.  Technicon Industrial Systems.   "Individual/Simultaneous Determination
    of Nitrogen and/or Phosphorus  in BD  Acid  Digests."  Industrial
    Method No. 320-7** W/B, Technicon Industrial Systems; Tarrytown, New
    York.  9 P. (1977).
9.  Jirka, A. M., Carter, M. J.,  May, D., and Fuller, F. D.  "Ultraview
    Semi-Automated Method for the Simultaneous Determination of Total
    Phosphorus and Total Kjeldahl Nitrogen in Wastewater."   Environ-
    mental Protection Agency, Central Regional Laboratory;  Chicago,
    Illinois.  27 p. (no date).
                                 3-206

-------
                              PHOSPHATES
                  (Soluble Reactive, Total,  Organic)

            Phosphates in the environment are known to exist  in  several
different chemical forms such as orthophosphate,  condensed  phosphates,
and organic phosphates.  The distinction "between  these forms  is  opera-
tionally defined but the conditions have been selected so they may be
                               i*
used for interpretive purposes.     A common  feature of each fraction is
that the phosphate is converted to orthophosphate,  which is then quanti-
fied using a colorimetric method.
            Soluble reactive phosphorus is defined  as  that  phosphate that
will pass through a O.ii5-y pore size membrane filter and react with the
colorimetric reagents without additional treatment.  Organic  phosphate
is defined as the difference between total phosphate and acid hydro-
lyzable phosphate.  The acid hydrolyzable phosphate fraction, in turn,
is defined as the phosphate concentration that results from the  acid
digestion of the sample at 100C.   The total phosphate fraction  is
determined by a strong acid digestion of the sample at elevated  tempera-
tures.
            Each of the colorimetric procedures for phosphate relies on
the formation of molybdophosphoric acid.  With two  of  the methods either
ascorbic acid  or stannous chloride1'2 is used to reduce the  heteropoly
acid to molybdenum blue, which is proportional to the  initial phosphate
concentration.  The third method relies on the formation of a yellow
complex when vanadium is added to molybdophosphoric acid.   The intensity
of the yellow complex is proportional to the initial phosphate concentra-
tion.1  The ascorbic acid and stannous chloride procedures  are subject to
arsenic interference and the vanadium procedure is  the least  sensitive
of the three procedures.

                      Sample Handling and Storage

            A flowchart for collection of phosphate samples is presented
*  References can be found on page 3-235.

                                3-207

-------
in Figures 3-29 to 3-31.   Sediments to be analyzed for  total  phosphates
may be stored in a wet, dried,  or frozen condition.  Work with raw and
treated sewage demonstrated that total phosphate was stable  for  k weeks
in acid-preserved samples  regardless of storage temperature.3  However,
if the various phosphate forms  are to be determined, the samples should
be processed as soon as possible and preferably within 1 day.
            Water samples  should be filtered immediately when  soluble
reactive phosphate is to be determined.  The sample may  be preserved
by freezing or the addition of  kO mg HgCl2/&.   (The addition of  HgCla
may interfere with phosphate analysis if the chloride concentration of
the sample is less than 50 mg/.4)  The addition of acid is  not  recom-
mended for the preservation of  soluble reactive phosphate since  it may
affect the orthophosphate-hydrolyzable phosphate equilibrium.  Separate
samples for hydrolyzable phosphate may be preserved with sulfuric acid.
Glass may be the preferred sample containers at low soluble  reactive
phosphate concentrations to minimize the effects of phosphate  adsorption.
                                 3-208

-------
            Figure  3-29.     Handling and storage of samples for orthophosphate  analysis

CORE SAMPLE
i

WATER SAMPLE 1>REDGE SMAPLE CORE SECTION
i * i
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ACIDIFY FILTER N0 T*^ENT STORE WET
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U) (Wl) (W2) (S1A) (SID)
1
ro
vo
SAMPLE DESIGNATION
PURPOSE
CONTAINER
SAMPLE TREATMENT
PRESERVATIVE
STORAGE TIME
DIGESTION SOLUTION
Wl W2 W3 S1A SIC SID
Total Water Soluble Used in Mobile Bloavail- Total
Cone. Water Elutriate Cone. ability Sediment
Cone. Cone.
G,P G,P G,P G,P G.P G,P
None Filter None None None None
4C >tC None AC 4C .C
(Minimize Air Contact. Keep Field Moist.)
2A hrs 2A hrs <1 w <1 w <1 w 
-------
         Figure  3-30.     Handling and  storage of samples for organic  phosphate analysis



( p
ACIDIFY
1
STORE
1
DIGEST
1
ANALYZE
LO (Wl)
ro
WATER SAMPLE CORE SAMPLE
I * *

FILTER NO TREATMENT ^REDGE SAMpLE CORE SECT|QN
1
i i
4r*" *
ACIDIFY STORE WET DRY
1 , 1
STORE STORE
1


DIGEST (r ELUTRIATE DIGEST DIGEST |
11 11
ANALYZE ANALYZE ANALYZE ANALYZE
(W2) (S1A) (SID) (S2)




^ir
FREEZE
1
STORE
1 ,
DIGEST
1
ANALYZE
(S3)

 SAMPLE DESIGNATION
PURPOSE


CONTAINER
SAMPLE TREATMENT
PRESERVATIVE

STORAGE TIME








DIGESTION SOLUTION


Wl W2 W3 S1A SID S2
Total Water Soluble Used in Mobile Total Total
S3
Total
Cone. Water Elutriate Cone. Sediment Sediment Sediment
Cone. Cone. Cone.
G,P G,P G,P G,P G,P G,P
Cone.
G,P
None Filter None None None Air Dry Freeze
H2SO,, H2SOft None 4C i|0C None
pH2 pH2 (Minimize Air Contact. Keep Field Moist.)
2k hr Zk hr <1 w <1 w <1 w
HOT HOT W3 HOT HOT
H2SOt, H2SOi| HzSOu H2SOt
None

-
HOT
H2SOi,
SAMPLE VOLUME OR WEIGHT
                       100 ml
                                 100 ml     Variable
                                                     100 ml
                                                                                   0.5-5 g    0.5-5 9    0.5-5 9

-------
                                                                                    CORE SAMPLE

J
*
WATER SAMPLE ' DREDGE SAMPLE CORE SECTION
t * J

ACIDIFY FILTER N0 "ENT STORE WET DRY
1 1 	 1
STORE ACIDIFY STORE
i 1


DIGEST STORE  ELUTRIATE "'"""I OIGEST DIGEST
i 1 1 	 1 1
ANALYZE ANALYZE ANALYZE ANALYZE ANALYZE
(Wl) (W2) (S1A) (SID) (S2)

I SAMPLE DESIGNATION
H
H PURPOSE
CONTAINER
SAMPLE TREATMENT
PRESERVATIVE
Wl W2 W3 S1A SIC SID 52

r
FREEZE
, i
STORE
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DIGEST
,1
ANALYZE
(S3)

S3


Total Water Soluble Used In Mobile Bioavall- Total Total Total
Cone. Water Elutriate Cone. ability Sediment Sediment Sediment
Cone. Cone. Cone. Cone.
G,P G,P G,P G,P G,P G,P G,P G,P
None Filter None None None None Air Dry Freeze
H2SOi, to H2SOu to None 4C 4C 
-------
                               PHOSPHATES
                          (Soluble Reactive)
                Procedures for Water Samples (W2,  SLA.)
Method 1:  Ascorbic Acid, Manual1
            Five colorimetric procedures are presented for the deter-
mination of phosphate.  Each are based on essentially the same principle.
However, the ascorbic acid and stannous chloride procedures are generally
more sensitive than the vanadomolybdate procedure.  The ascorbic acid
methodology has probably received wider usage because of relative ease
and stability of the reducing agent.
Apparatus
Colorimeter or spectrophotometer equipped for use  at 885 nm.   A light
      path of 2. 5 cm or longer would be preferred
Acid-washed glassware:  all glassware should be washed in hot 1:1 HC1 and
      rinsed with distilled water.  Commercial detergents should never
      be used
Reagents
Sulfuric acid solution, 5 N:  dilute 70 ml cone. H2S04 to 500 ml with
      distilled water.
Potassium antimonyl tartrate:  dissolve 1.3715 g KtSbOjCitH^Oe   1/2 H20
      in kOO ml distilled water.  Dilute to 500 ml with distilled water
      and store in a glass-stoppered bottle.
Ammonium molybdate solution:  dissolve 20 g (NHi* )eMo702it '  ^ H20 in 500 ml
      distilled water.  Store in a plastic bottle at hC.
Ascorbic acid:  dissolve 1.76 g ascorbic acid in 100 ml distilled water.
      The solution is stable for about 1 week at hC.
Combined reagent:  mix the above reagents in the following  proportions
      for 100 ml combined reagent:  50 ml 5 N. H2SOit, 5 ml potassium
      antimonyl tartrate, 15 ml ammonium molybdate, and 30  ml ascorbic
      acid.   All reagents must reach room temperature before they are
      mixed.  Add the reagents in the order given and mix the resultant
      solution after each addition.  The combined reagent is stable for
      h hr.
Stock phosphate solution:  dissolve 219.5 mg anhydrous potassium dihydro-
      gen phosphate in distilled water and dilute to 1000 ml.  1.0 ml =
      50.0 yg POi,-P.
Procedure
            Pipet 50 ml W2 or S1A sample or standard into a 125-ml
Brlenmeyer flask.  Add 1 drop phenolphthalein indicator. If a red color
develops, add 5 N_ H2SOi* dropwise until the color disappears.  Add 8.0 ml
                                 3-212

-------
combined reagent and mix.
            After 10 min, but prior to 30 min after adding the combined
reagent, measure the absorbance of each sample at 885 nm relative to the
reagent blank.
            If the samples are turbid or colored, a blank must be prepared
by adding all reagents except ascorbic acid and potassium antimonyl tar-
trate to the sample.  Measure the absorbance relative to the reagent
blank and subtract from each sample.
Calculations
            Prepare a calibration curve by plotting known phosphate
concentration vs. standard absorbance.  Determine the phosphate concen-
trations of the samples by comparing the measured sample absorbance with
the standard curve.

Method 2:  Ascorbic Acid, Automated4
Apparatus
Technicon autoanalyzer system consisting of:
      a_.  Sampler
      b_.  Manifold (AAl) or Analytical Cartridge (AAIl)
      c_.  Proportioning pump
      d_.  Heating bath, 50C
      e_.  Colorimeter equipped with 15- or 50-mm tubular flow cell
      f_.  650- to 660- or 880-nm filter
      .  Recorder
      h_.  Digital printer for AAl I (optional)
Acid-washed glassware:  all glassware should be washed in hot 1:1 HC1
      and rinsed in distilled water.  Commercial detergents should never
      be used.
Reagents
Sulfuric a'cid solution:  dilute TO ml cone. HaSOu to 500 ml with distilled
      water.
Potassium antimonyl tartrate:  dissolve 0.3 g K(SbO)CitH1|06  1/2 H20
      in distilled water and dilute to 100 ml.  Store at ^C in a dark,
      glass-stoppered bottle.
Ammonium molybdate:  dissolve h g (NHi,) eMoyOai*' U H20 in 100 ml distilled
      water.  Store in a plastic bottle at UC.
                                 3-213

-------
Ascorbic acid:  dissolve 1.8 g ascorbic acid in 100 ml distilled water.
      The solution is stable for about 1 week when stored at UC.
Combined reagent:  mix the following reagents in the following propor-
      tions and order to prepare 100 ml of reagent:  50 ml sulfuric
      acid, 5 ml potassium antimonyl tartrate, 15 ml ammonium molybdate,
      and 30 ml ascorbic acid.  All reagents must reach room temperature
      before they are mixed.  If turbidity forms in the combined reagent,
      shake and let it stand for a few minutes until the turbidity
      disappears.  This solution is stable for approximately k hr and
      must be prepared fresh for each run.
Stock phosphate solution:  dissolve 0.^393 g predried (105C for 1 hr)
      KH2POi, in distilled water and dilute to 1000 ml.  1.0 ml = 0.1 mg P.
Procedure
            To 50 ml of W2 or S1A sample, add 1 drop phenolphthalein
indicator.  If a red color develops, add sulfuric acid solution dropwise
to discharge the color.  Acid samples must be neutralized with 1 N_ NaOH.
            Set up the autoanalyzer manifold as shown in Figure 3-32 or
3-33.  Allow the colorimeter and recorder 30 min to warm up.  Obtain a
stable baseline by using all reagents and feeding distilled water through
the sample line.
            Place standards in sample tray in order of decreasing con-
centration.  Complete tray with unknown samples.  Switch sample line from
distilled water to sampler and begin analyses.
Calculations
            Prepare a standard curve by plotting peak height vs. standard
concentration.  Compute sample concentrations by comparing peak sample
height with the standard curve.

Method 3:  Stannous Chloride, Manual1
Apparatus
Colorimeter or spectrophotometer equipped for use at 690 nm
Acid-washed glassware:  all glassware used in the procedure should be
      washed in hot 1:1 HC1 and rinsed thoroughly with distilled water.
      Commercial detergents should not be used
Reagents
Sulfuric acid solution:  dilute 300 ml cone. HaSOit to approximately
      600 ml with distilled water.  When solution has cooled, add U ml
      cone. HNOs and dilute to 1 .
                                 3-211*

-------
                   Figure 3-32.    AAI manifold for the ascorbic acid determination of phosphorus
u>
     SM
     LM
       50C
       HEATING
        BATH

fixing Co
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OC

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O.A2 MIXED
REAGENT
2.00 WASTE
.
                 COLORIMETER

                  50 mm FLOW CELL
                  650-660 or 880 nm FILTER
RECORDER

-------
              Figure 3-33.  AAII cartridge for the ascorbic acid determination of phosphorus
ro
WASH WATER
TO SAMPLER

0000
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         WASTE
                                     RECORDER
                                                       DIGITAL
                                                        PRINTER
                       COLORIMETER
                        50 mm FLOW CELL
                        650-660 or 880 nm FILTER

-------
Annnonium molybdate reagent I:  dissolve 25 g (NHi, )6Mo702it '  ^ Had in
      175 ml distilled water.  Cautiously add 280 ml cone.  HaSOt*  to hOO ml
      distilled vater.  Cool, add the molybdate solution, and dilute to
      1  with distilled water.
Stannous chloride reagent I:  dissolve 2.5 g fresh SnCla    2 HaO in 100 ml
      glycerol.  Heat in a water "bath and stir with a glass rod to hasten
      dissolution.
Standard phosphate solution:  dissolve 219-5 mg anhydrous potassium
      dihydrogen phosphate, KHaPOij ,  and dilute to 1000 ml with distilled
      water.  1.0 ml = 50 yg POt,-P/.
Procedure
            To 100 ml of W2 or S1A sample or standard containing not
more than 0.2 mg P and free from color and turbidity, add 1 drop phenol-
phthalein indicator.  If the sample turns pink, add sulfuric acid drop-
wise until the color disappears.  If more than 0.25 ml is required, con-
tinue titrating until the color disappears but dilute an aliquot of the
pH adjusted sample to 100 ml with distilled water for analysis.
            The rate of color development and the color intensity of
the final solution are dependent upon temperature.  Therefore, samples
and standards should be equilibrated at the same temperature and be
within 2C of each other at the time of color development.
            Add U.O ml molybdate reagent I to the pH-adjusted sample and
mix thoroughly.  Add 0.5 ml (10 drops) stannous chloride and mix
thoroughly.  Ten minutes after the addition of the color imetric reagents,
but prior to 12 min after the addition of the reagents, determine the
absorbance of the standard or sample relative to a distilled water blank
at 690 ran.
NOTE:  Because of the dependence of color intensity on time, a serious
       effort must be made to adhere to a strict time schedule and
       allow a constant color development time for all standards and
       samples .
Calculations
            Prepare a standard curve by plotting observed standard
absorbance vs. phosphate concentration.  Compare observed sample
absorbance with the standard curve to determine sample phosphate
concentration.
                                 3-217

-------
Method h:  Stannous Chloride, Automated
Apparatus
Technicon autoanalyzer system (Figure 3-3^ or 3-35) consisting of:
      a_.  Sampler
      b.  Heating bath, 30 C
      c_.  Manifold (AAl) or Analytical Cartridge (AAIl)
      d_.  Proportioning pump
      e_.  Colorimeter, 50-mm flow cells, 660- nm filters
      f_.  Recorder
      g_.  Range expander
Reagents
Ammonium molybdate solution:  dissolve 25 g ammonium molybdate,
      024   J* H20, in 175 ml distilled water.  Dilute 155 ml cone.  H2SOi
      to kOO ml with distilled water.  Mix the two solutions and dilute
      to 1 .
Stock stannous chloride:  dissolve 5 g stannous chloride, SnCla '  2
      in 25 ml cone. HC1 and dilute to 500 ml with distilled water.  This
      solution is stable for approximately 2 weeks at 5C.
Stannous chloride working solution:  mix 30 ml stock stannous chloride
      solution with 25 ml cone. HC1 and dilute to 500 ml with distilled
      water.  This solution is stable for approximately 12 hr .
Sulfuric acid solution:  add 300 ml cone. H2SOi| to distilled water and
      dilute to 1 I.
Standard phosphate solution:  dissolve 219-5 mg anhydrous potassium
      dihydrogen phosphate, KHaPOit, and dilute to 1000 ml with distilled
      water.  1.0 ml = 50 ug P04-P.
Prepare appropriate working phosphate solutions by diluting the standard
      phosphate solution.  Working solutions should be made daily.
Procedure
            Set up the autoanalyzer manifold as shown in Figure 3-3^ or
Figure 3-35-  Figure 3-3^ is for samples in the phosphate range of 0 to
50 yg/A and Figure 3-35 is for samples in the phosphate range of 50 to
500 ug/.
            Allow the colorimeter and recorder 30 min to warm up.   Obtain
a stable baseline by using all reagents and feeding distilled water through
the sample line.
            Place standards in the sample tray in order of decreasing
concentration.  Fill the remainder of the tray with W2 or S1A samples.

                                 3-218

-------
              Figure 3-31*.   Stannous chloride  manifold for the determination of phosphorus,
                                          low  level (0-50 yg/A)
(jO
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AIR
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WASTE FR

                                            PROPORTIONING
                                                PUMP
                              RANGE
                             EXPANDER
RECORDER
             COLORIMETER
              50 mm TUBULAR f/c
              660 mu FILTERS
                                                                                                  SAMPLER
                                                                                                   20/hr

-------
                Figure 3-35.  Stannous chloride manifold for the determination of phosphorus,
                                            high level (50-500 yg/)
     H3
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2.5 WASTE FROM COLORIMETER
'
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             COLORIMETER
              50 mm TUBULAR f/c
              660 mu FILTERS

-------
Switch sample line from distilled water to sampler and begin analysis.
Wash water should be acidified with sulfuric acid at the rate of 10 ml
30 percent HzSO^/H distilled water.
            Every time a new batch of stannous chloride reagent is used,
a new set of phosphate standards must be run.  Standard solutions
should be run periodically to check the validity of the calibration
curve.
Calculations
            Prepare a standard curve by plotting peak height vs.
standard concentration.  Compute sample concentrations by comparing
sample peak height with the standard curve.

Method 5:  Vanadomolybdophosphoric Acid, Manual1
Apparatus
Spectrophotometer equipped with a ^70- to l*90-nm filter
Glassware:  all glassware should be washed in hot 1:1 HC1 and rinsed
      thoroughly with distilled water.  Commercial detergents should
      not be used
Reagents
Activated carbon.
Vanadate-molybdate reagent:
      Solution A:  dissolve 25 g ammonium molybdate, (NHiJ 6Mo702i '  ^ H20,
                   in UOO ml distilled water.
      Solution B:  dissolve 1.25 g ammonium metavanadate, NH^VOa, in
                   300 ml boiling distilled water.  Cool and add 330 ml
                   cone. HC1.
      Cool Solution B to room temperature, add Solution A, and dilute
      to 1 & with distilled water.
Standard phosphate solution:  dissolve 219-5 Dig anhydrous potassium
      dihydrogen phosphate,  KHaPOt,, in distilled water and dilute to 1 
      1.0 ml = 50 yg POij-P.
Procedure
            Remove any excessive color from the sample by shaking 50 ml
sample with 200 mg activated carbon for 5 min.  Remove the carbon by
filtering the sample through a Whatman No. U2 filter paper or the
equivalent.
NOTE:  Check each batch of activated carbon for phosphate release.
                                 3-221

-------
            Transfer 35 ml of phosphate standard or sample,  containing
1000 ug P or less, to a 50-ml volumetric flask.   Add 10 ml vanadate-
molybdate reagent and dilute to volume with distilled water.   Prepare a
reagent blank using 35 ml distilled water and 10 ml vanadate-molybdate
reagent.
            Allow 10 min for color development and measure the absorbance
of the sample vs. the reagent blank.  The wavelength to be used depends
on the sensitivity desired.  A wavelength of 1+70 nm is usually used but
wavelengths between kOO and ^90 nm can be used.
Calculations
            Prepare a calibration curve by plotting standard absorbance
vs. phosphate concentration.  Determine the phosphate concentration of
the samples by comparing sample absorbance with the standard curve.
                                 3-222

-------
                  Procedure for Sediment Samples (SID)

            This procedure is operationally defined.  Total soluble
phosphate is defined as the amount of phosphate leached from sediment
in a stated time period.  It is recommended that moist sediments be
used for this determination.  The reason for this restriction is that
drying or freezing of sediment samples may cause irreversible conversion
or sorption of the phosphate in the sediment sample (Figure 3-29).  The
final leachate is then analyzed for soluble phosphate using one of the
procedures provided in the section on water analysis.
Sample preparation5
            Place 10 g of blended, wet sediment (SID) into a 100-ml
beaker.  Add 50 ml distilled water and mix.  Allow the suspension to
settle overnight.
            Centrifuge, if necessary, then filter through a 0.^5^
pore-size membrane filter.  Do not wash the filter.  Transfer the
filtrate to a 250~ml beaker.  Add 1 ml strong acid solution (300 ml
cone. H2S04 and h ml cone. HN03 diluted to 1 &) and O.k g potassium
persulfate (IC,S208 ).  Boil solution for 90 min, adding distilled
water, if necessary, to keep the volume between 25 and 50 ml.
            Cool the sample, add 1 drop phenolphthalein, and titrate
with 1 IT NaOH to a faint pink color.  Transfer to a 100-ml volumetric
flask and dilute to volume with distilled water.
Quantification procedure
            Analyze the digested leachate by one of the five methods
presented in the section for soluble reactive phosphate analysis in
water samples.
Calculations
            Compare the absorbance of the sediment leachate with a
standard phosphate curve to determine the phosphate concentration.  The
soluble phosphate concentration of the sediment sample is calculated as
                                 3-223

-------
follows :
        Soluble Phosphate mg/kg (vet basis) = (x) (0-1(j (1000)
                                                    1
        Soluble Phosphate mg/kg (dry basis) =  j

where
       x = phosphate concentration in leachate ,  mg POit-P/
     0.1 = volume of leachate,  (0.1 & as written)
       g = wet weight of sample used, g
     % S = percent solids in sediment sample as a decimal fraction

-------
                               PHOSPHATES

                                (Total)


              Procedures for Water Samples (Wl, W2, S1A)


            There are three digestion methods available to determine

total phosphate.  In terms of increasing severity, these are:  (a)

persulfate digestion, (b) sulfuric acid-nitric acid digestion, and

(c) perchloric acid digestion.  A less severe digestion method should
only be used when it has been shown to be equivalent to the most

severe method for the type of samples being analyzed.

            Ea.ch of the procedures subjects the water samples to a
strong acid digestion at elevated temperatures.  The digests are then

filtered to remove remaining particulate matter and diluted to a con-

venient volume.  Analyze the digests for phosphate using one of the

procedures in the soluble reactive phosphate section and report the

results as total phosphate.

Sample preparation

            Select a digestion procedure from those indicated below

and proceed as indicated.

            a.  Persulfate digestion.*'2 Transfer a 50-ml aliquot of
                Wl, W2, or S1A sample or standard to a 125-ml Erlenmeyer
                flask.  Add 1 drop of phenolphthalein indicator solution.
                If a red color develops, add 10.8 IJ HzSOi, dropwise until
                the color disappears.  Add 1.0 ml of the sulfuric acid
                solution (300 ml cone. RzSOn diluted to 1 &) and O.U g
                solid ammonium persulfate.
                Either (l) boil the acidified sample on a preheated hot
                plate for hO min or until the volume is reduced to 10 ml,
                or (2) autoclave the sample for 30 min at 1.0 to l.U
                kg/cm2 (15 to 20 psig).  Cool the samples, neutralize
                to the phenolphthalein endpoint with 1 N_ NaOH, and
                dilute to 100 ml with distilled water.
            b_.  Sulfuric acid-iiitric acid digestion.1  Transfer a 50-ml
                aliquot of Wl, W2, or S1A sample or standard to a micro
                Kjeldahl flask.  Add 1 ml cone. HzSOij and 5 ml cone.
                HN03-  Digest on a micro-Kjeldahl digestion rack to a
                voluiae of 1 ml and continue heating until HKOs is removed
                and solution is colorless.
                                3-225

-------
                Cool and add 20 ml distilled water.   Add 1 drop phenol-
                phthalein and neutralize with 1 N_ NaOH to a faint pink
                color.   Filter, if necessary, and transfer the digest
                to a 100-ml volumetric flask.  Wash  the digestion flask
                and filter into the volumetric flask.  Dilute to 100 ml
                with distilled water.

                Perchloric acid digestion.    Transfer a 50-ml aliquot
                of Wl,  W2, or S1A sample or standard to a 125-ml
                Erlenmeyer flask.  Acidify to the methyl orange
                endpoint with cone. HNOa.  Add an excess of 5 ml cone.
                HNOa.  Evaporate the sample to 15 to 20 ml on a steam
                "bath or hot plate.  Cover the sample with a watch glass
                when necessary to avoid sample loss  due to splattering.

                Add 10 ml cone. HN03,  10 ml JO to 72 percent HCIO^,  and
                a few boiling chips.  Heat gently on a hot plate until
                the evolution of dense white HClOit fumes.  If the
                solution is not clear, add 10 ml cone. HN03 and continue
                heating.
                Cool the solution and neutralize with 6 N_ NaOH to the
                phenolphthalein endpoint.  Filter the solution, if
                necessary, and dilute to 100 ml with distilled water.
 lantification procedure
            The digestion procedure should produce a homogeneous,  liquid

phase digest.  These samples should be analyzed for phosphate using one

of the procedures for soluble reactive phosphate in water samples.

Calculations
            Determine the phosphate concentration of the digests by

comparing measured sample absorbances with a standard phosphate curve.

Calculate the phosphate concentration of the initial sample by multi-

plying the phosphate concentration of the digest by the ratio of digest

volume to initial sample volume:

                     Total phosphate mg/ = lililL
                                               s

where

     x = phosphate concentration in digest, mg/

     d = volume of digest, ml (100 ml as written)

     s = sample volume, ml (50 ml as written)

            Report the results for W2 and S1A samples as total soluble

phosphate.  Report the results for Wl samples as total phosphate.
                                3-226

-------
             Procedures for Sediment Samples (SID, S2, S3)

            There are numerous methods available for the digestion of
sediment samples to be analyzed for phosphate.  In fact, Aspila et al.6
reported a literature review summarizing 77 methods.  Most procedures
consist of strong acid digestion or treatment with an oxidizing agent
and a strong acid.
            A common feature of the digestion procedures is that the
sample treatments are designed to convert all the phosphate compounds
to ortho phosphate.  The orthophosphate is then quantified using one
of the analytical procedures presented earlier.  Therefore, a choice
of digestion techniques is .-presented but the analytical procedures are
not repeated.

                      Sample Handling and Storage

            Phosphate may interconvert between several forms.  However,
the compounds are not considered volatile.  Therefore, if it is decided
to run total phosphate, samples may be stored in a field moist, dried,
or frozen condition.  This information is summarized in Figure 3-31.
Sample pretreatment
            Weigh out a 0.5- to 1.0-g dry weight equivalent of the
sediment sample.  Continue with one of the digestion procedures
presented in a_ through e_ below:
            a_.   Perchloric acid digestion.*1'7  Weigh into a 125-ml bea-
                ker an 0.5-g dry weight equivalent of sediment sample.
                Add 25 ml distilled water and 5 ml cone.  HNO;j.
                Mix the sample and evaporate on a hot plate to 5 to
                10 ml.  Transfer the sample to a 125-ml conical flask.
                Rinse the beaker with 5 ml cone. HNOs.  Add 5 ml cone.
                HN03, 10 ml of 70 to 72 percent HClOit, and a few
                boiling stones.  Heat on a hot plate and evaporate to
* Stone or asbestos cement hoods recommended when perchloric acid is to
  be used.
                                3-227

-------
    the evolution of dense white HC10i fumes.  If the
    solution is not clear at this point, cover the flask
    with a watch glass and continue heating until it clears.
    Add additional cone. HNOa, if necessary.

    Cool the solution and neutralize to the phenolphthalein
    endpoint with 6 H_ WaOH.

    Filter the sample, if necessary; transfer to a volumetric
    flask, and dilute to volume.  Standards should be carried
    through the digestion procedure to correct for an ionic
    strength effect on the colorimetric procedure.
b_.  Sulfuric acid-nitric acid digestion.5  Weigh out 0.5 to
    1.0 g sediment sample.  Transfer to a 250-ml Erlenmeyer
    flask using a minimum amount of water.  Add 5 ml cone.
    HaSOi* and 25 ml cone. HNOs.  Mix the suspension well
    after the addition of each acid.

    Digest slowly on a hot plate with medium heat.  Avoid
    use of high heat as this may cause superheating and
    result in sample loss.  Continue heating until the
    evolution of white fumes.  If the sample is not clear,
    add cone. HNOs and continue the digestion process until
    a clear digestate is obtained.
    NOTE:  Do not heat sample to dryness.

    Add 25 to 50 ml distilled water to the hot sample and
    filter immediately.  Pour the filtrate through a H- by
    3A-in. column of cation exchange resin to remove iron.
    Wash the column with approximately 25 ml distilled water.
    Titrate the sample with 6 N_ NaOH to the phenolphthalein
    endpoint.  Back titrate the sample dropwise with 1 IT
    H^SOit until colorless.  Transfer the solution to a
    100-ml volumetric flask and dilute to volume.

c_.  Persulfate digestion (sealed bomb).6  Accurately weigh
    a 0.3- to 0.5-g dry weight equivalent of the sample and
    transfer to a Parr PFTE bomb (Parr Instrument Co.,
    Moline, Illinois).  Add 3.0 +_ 0.1 g potassium persulfate
    and 5-0 ml cone. HaSO-*.  Seal the bomb and heat at
    135C for 2 hr.
    Transfer the contents of the bomb to a 500-ml volumetric
    flask and dilute with distilled water.
d.  Persulfate digestion.6  Weigh 0.5-g dry weight equivalent
    of the sample and transfer to a 150-ml beaker.  Add 10 ml
    30 percent HaSOij and 2 g potassium persulfate.  Mix the
    suspension and heat on a hot plate for 1 hr.  Filter, if
    necessary, into a 100-ml volumetric flask and dilute
    to volume.
                     3-228

-------
                e .   Block digestion.8 ' 9  Weigh 0 . 5 g dry weight equiva-
                    lent of sediment and transfer to a Technicon glass
                    digestion tube (No. 11^-002^-02).  Add 10 ml
                    digestion solution, 1 ml HgO solution, and 2 to 3
                    boiling stones (ChemPlast, Inc., Wayne, New Jersey).
                    Mix samples using a vortex mixer.

                    NOTE:  The digestion solution is prepared by gradu-
                           ally adding 600-g KaSOi, to 1-& cone . HaSOi* .
                           Allow solution to cool to room temperature,
                           stopper, and store above 20 C.

                    NOTE:  The mercury solution is prepared by
                           dissolving 5-g HgO in 100 ml 10 percent
                    Place samples in digestion rack (Technicon No.
                    0009-02) and place the rack on a block digester
                    (Technicon No. BD-AO).  Heat samples 1 hr at 200C,
                    followed by 1 hr at 3TOC.  Cool samples and
                    dilute to 75 ml.
Quantification procedure

            Dilute the sediment digests to a convenient volume  Analyze
the digests for phosphate using one of the procedures described earlier.

Calculations
            Determine the phosphate concentrations of the digests by

comparing sample absorbance with the standard phosphate curve.  Calcu-
late the phosphate concentration of the initial sediment samples as

follows :

           Total phosphate mg/kg (wet weight) = -
           Total phosphate mg/kg (dry weight) =  */ V la g\

where
       x = phosphate concentration in sediment digest,  mg/Jl
       y = final volume of sediment digest ,  H

       g = wet weight of sample digest, g
     % S = percent solids in sediment sample as a decimal fraction
                                3-229

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                               PHOSPHATES
                               (Organic)

            The determination of organic phosphate requires the analyses
of two samples.  One sample is subjected to a total phosphate digestion
and a second sample is subjected to an acid hydrolysis.  The digests are
analyzed for phosphate using one of the colorimetric procedures and
organic phosphate is calculated by subtracting the acid hydrolysis
results from the total phosphate results.

                      Sample Handling and Storage

            Samples for organic phosphate analysis may be stored in
either glass or plastic containers and preserved with the addition of
sulfuric acid.   However, since the maximum recommended holding time for
hydrolyzable phosphate and total phosphate is presently 2k hr ,** the
holding time for organic phosphate samples should also be 2k hr
(Figure 3-30).

              Procedures for Water Samples (Wl, W2, S1A)1

Sample _prep_ar_atip_n
            Two subsamples must be prepared for phosphate analvsis in
order to determine organic phosphate.  One sample is digested in strong
acid at elevated temperatures.  A second sample is hydrolyzed under less
severe conditions.
            i.   Total digest.  Transfer a 50-ml sample to an Erlenmeyer
                flask or a micro Kjeldahl flask and proceed with either
                the persulfate digestion, sulfuric acid-nitric acid
                digestion, or perchloric acid digestion, as described
                for the total phosphate analysis of water samples.
            b.   To a second 100-ml sample, or a sample aliquot diluted
                to 100 ml, add 1 drop phenolphthalein indicator
                solution.  If a red color develops, add a strong acid
                solution dropwise until the color disappears.  Add a
                1 ml excess of strong acid.
                Boil the solution for 90 min.  Add distilled water as
                                3-230

-------
                    required to keep the water level between 25 and
                    50 ml.  Cool the sample, neutralize with sodium
                    hydroxide to a faint pink color, and dilute to 100 ml
                    with distilled water.  As an alternative, the sample
                    may be autoclaved for 30 min at 1.0 to l.U kg/cm
                    (15 to 20 psig).
                    Prepare a series of orthophosphate standards and
                    process through the hydrolysis procedure.  This is
                    necessary to compensate for the ionic strength
                    effects due to the hydrolysis procedure.  Cool,
                    neutralize, and dilute the standards as indicated
                    with the samples.
Quantification procedure
            Analyze the digests from both procedure a_ and procedure b_
for orthophosphate using one of the procedures provided in the section
for soluble reactive phosphate in water samples.  Label the digests
from procedure a_ as total phosphate and the digests from procedure b_
as hydrolyzable phosphate.
Calculations
            Prepare a standard phosphate curve by plotting absorbance
vs. phosphate concentration.  Determine the phosphate concentration of
the sample digests by comparing the sample absorbance with the standard
curve.  Correct the digest phosphate concentration for any dilution
during the sample pretreatment:
                       Sample Phosphate = -^1111
                                             s
where
     x = the phosphate concentration of the digest, ug/&
     d = final volume of sample digest, ml
     s = initial sample volume, ml
            The organic phosphate concentration is then calculated as:
     Organic Phosphate = Total Phosphate - Hydrolyzable Phosphate
            When a total water sample (Wl) is used, the result is total
organic phosphate.  When a filtered water sample (W2, S1A) is used, the
result is soluble organic phosphate or filterable organic phosphate.
                                3-231

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             Procedures for Sediment Samples (SID,  S2,  S3)

            Two procedures are presented for pretreatment of sediment
samples to determine organic phosphate.   Because of the difference in
techniques, organic phosphate should be considered an operationally
defined parameter.
Method 1:  Acid Hydrolysis1
Sample pretreatment
            It is necessary to prepare two subsamples for phosphate
analysis to calculate organic phosphate.
            a.  Total digest.  Weigh out a 0.3- to 1.0-g dry weight
                equivalent of the sediment sample and proceed as
                described in the section on total phosphate analysis
                of sediments.
            b_.  Place 5 to 10 g sediment in a 150-ml beaker.  Add
                50 ml distilled water and 5 ml strong acid solution
                (300 ml cone. H2SOi+ and It ml cone.  HN03 diluted to 1 &).
                Mix the suspension and boil on a hot plate for 90 min.
                Add distilled water as necessary to maintain a liquid
                level of 25 to 50 ml.
                Cool the sample and filter.  Add 1 drop of phenol-
                phthalein indicator and neutralize to a faint pink
                color with 1 IT NaOH.  Dilute the sample to 100 ml with
                distilled water.
                Prepare a series of orthophosphate standards and
                process through the hydrolysis procedure.  This is
                necessary to compensate for the ionic strength effects
                due to the hydrolysis procedure.  Cool, neutralize,
                and dilute the standards as indicated with the samples.
Quantification procedure
            Analyze the sediment digests and standards carried through
the digestion procedures for orthophosphate using one of the methods
provided for the analysis of soluble reactive phosphate in water.
Calculations
            Prepare the appropriate standard curves by plotting standard
absorbance vs. phosphate concentration.   Determine the phosphate
                                 3-232

-------
concentration of the sediment digests by comparing sample absorbance with
the standard curve.  Calculate the total and hydrolyzable phosphate con-
centrations of the sediment sample as:
       Phosphate Concentration mg/kg  (wet weight) = 	-,v	

       Phosphate Concentration mg/kg  (dry weight) = -,w^ _,N
                                                      (g) (7 S)
where
       x = phosphate concentration of the sediment digest, mg/&
       v = volume of sample digest, H
       g = wet weight of sediment sample, g
     % S = percent solids of sediment (written as a decimal fraction)
The results from pretreatment a_ are total phosphate concentrations and
the results from pretreatment b_ are hydrolyzable phosphate.  Sediment
organic phosphate concentrations are calculated as total phosphate minus
hydrolyzable phosphate.

Method 2:  Acid Extraction6
Sample  retreatment
            It is necessary to prepare two subsamples for phosphate
analysis to calculate organic phosphate.
            &.  Total digest.  Weigh out a 0.3- to 1.0-g dry weight
                equivalent of the sediment sample and proceed with one
                of the five digestion procedures described in the
                section on total phosphate analysis of sediments.
          -  b_.  Acid extract..  Weigh a second 0.3- to 0.5-g dry weight
                sample and transfer to a 100-ml volumetric flask.
                Extract the sample with 50 ml 1 N_ HC1 for l6 hr in a
                room temperature water bath.  Neutralize the sample
                to the phenolphthalein endpoint with 6 IT NaOH.
Quantification procedure
            Dilute the total digest from a_ and the acid extract from b_
to convenient volumes.  Analyze for orthophosphate using one of the
methods provided for the analysis of soluble reactive phosphate in water.
Calculations
            Prepare the appropriate standard curve by plotting standard
absorbance vs. phosphate concentration.   Determine the phosphate
                                 3-233

-------
concentration of the sediment digests by comparing sample absorbance with
the standard curve.  Calculate the total and acid extractable phosphate
concentrations of the sediment sample as:
       Phosphate concentration mg/kg (wet weight) = -T^-T	-

       Phosphate concentration mg/kg (dry weight) =   /  T/i _>-
                                                      (g) (f> o)
where
       x = phosphate concentration of the sediment digest, mg/&
       v = volume of sediment digest, &
       g = wet weight of sediment sample, g
     % S = percent solids of sediment (expressed as a decimal fraction)
The results from pretreatment a_ are total phosphate concentrations and
the results from pretreatment b_ are acid extractable phosphate.  Sediment
organic phosphate concentrations are calculated as total phosphate minus
acid extractable phosphate.

-------
                              References
1.  American Public Health Association.   Standard Methods for the
    Examination of Water and Wastewater.  l^th Edition.   APHA;
    New York, New York.   1193 p.  (1976).
2.  Environment Canada.   "Analytical Methods Manual."  Inland Water
    Directorate, Water Quality Branch;  Ottawa, Ontario  Canada (197*0 
3.  Carter, Mark J.  "Wastewater  Sample Preservation Data for Twenty
    Common Water Quality Parameters."  Chemistry Branch, National
    Enforcement Investigation Center, U.  S.  EPA; Denver, Colorado
    (1978).
U.  Environmental Protection Agency. "Manual of Methods for Chemical
    Analysis of Water and Wastes."  Methods  Development and Quality
    Assurance Research Laboratory, National  Environmental Research
    Center; Cincinnati,  Ohio.  298 p. (197*0-
5.  Great Lakes Region Committee  on Analytical Methods.  "Chemical
    Laboratory Manual for Bottom  Sediments."  U. S.  Department of the
    Interior, Great Lakes Region; Chicago, Illinois.  96 p. (1968).
6.  Aspila, K. I., Agemian, H., and Chan, A. S. Y.  "A  Semi-Automated
    Method for the Determination  of Inorganic, Organic  and Total
    Phosphate in Sediments."  Analyst 101:187-197 (1976).
7.  Andersen, J. M.  "An Ignition Method for Determination of Total
    Phosphorus in Lake Sediments."  Water Res. 10:329-331 (1976).
8.  Jirka, A. M., Carter, M. J.,  May, D. , and Fuller, F. D.  "Ultramicro
    Semiautomated Method for Simultaneous Determination of Total
    Phosphorus and Total Kjeldahl Nitrogen in Wastewater."  Env.  Sci.
    and Tech. 10:1038-10^ (1976).
9.  Anon.  "Individual/Simultaneous Determination of Nitrogen and/or
    Phosphorus in BD and Digests."  Technicon Industrial Systems
    Industrial Method No. 329-7U  W/B.  Technicon Industrial Systems;
    Tarrytown, New York.  9 p. (1977).
                                3-235

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                               SULFIDES

            Sulfid.es are the salts of a weak acid,  HaS.   Therefore,
depending on the pH, sulfides may exist as H2S,  HS  ,  or  S .   Sulfides
are of concern because they may "be potentially toxic  (toxicity will
vary with pH) and they may create unaesthetic conditions (odor of
rotten eggs.

                     Sampling Procedure and Storage

            The collection of samples for sulfide analysis presents
two problems.  The first is that H2S is a gas.  Consequently, sulfide
may be lost by volatilization and/or gas stripping.  The second
problem is that sulfides are reducing agents and can  be  oxidized by
dissolved oxygen.  When these factors were considered, the following
approach was developed for handling of sulfide samples (Figure 3-36).
Water samples should be collected with a minimum of turbulence as a
precaution against volatilization and oxidation of  sulfides.   Also,  a
zinc acetate preservative should be used to precipitate  zinc  sulfide.
This is accomplished by adding k drops 2 N_ zinc acetate  in a  100-ml
bottle, completely filling with sample, and tightly sealing the bottle.
If it is desired to determine soluble sulfides,  the sample should be
flocculated with alum to remove particulate matter.  Filtration is
likely to expose the sample to the atmosphere and cause  sulfide
oxidation.  Sulfide analyses should be completed as soon as possible
and preferably within 2k hr.
            Only wet sediment samples should be analyzed for  sulfides
because of the likelihood of sample oxidation during  the drying or the
freezing/thawing cycle.  Samples may be treated with  zinc acetate but
the efficiency of this step is unknown.  The storage  time for sediment
samples is not known.  Therefore, samples should be processed as soon
as possible.
                                3-236

-------
              Figure 3-36.     Handling  and storage  of samples for  sulfide analysis

1 CORE SAMPLE
*

WATER SAMPLE DREDGE SAMPLE CORE SECTION
1 * 1
1 *
ALUM
FLOCCULAT
STORE WET
ON
i
STORE STORE
i ' \ '

1
DISTILL
1
ANALYZE ANALYZE A/c^n?E
U) fWl) (W2) (S]0>
ro
"^ SAMPLE DESIGNATION
PURPOSE
CONTAINER
SAMPLE TREATMENT
PRESERVATIVE
STORAGE TIME
DIGESTION SOLUTION
Wl W2 SID
Total Water Soluble Total
Cone. Water Sediment
Cone. Cone.
G,P G,P G,P
None Filter None
Zinc Zinc <4C
Acetate Acetate (Zinc Acetate)
(Minimize Air Contact. Keep Field Moist.)
2k hr m hr As soon as possible.
Amlne- Amine- ^^1
Sulfur !c Sulfurlc
Acid Acid
SAMPLE VOLUME OR WEIGHT
                      25 ml
25 ml
                                                                                1-59

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                                                      A*

                Procedures for Water Samples (Wl, W2)1
Method 1:  Methylene Blue, Colorimetric
Apparatus

Matched test tubes, approximately 125 mm long and 15 mm O.D.
Droppers, delivering 20 drops/ml methylene blue solution.   To obtain
      uniform drops it is essential to hold the dropper in a  vertical
      position and to allow the drops to form slowly
Spectrophotometer or filter photometer for use at a wavelength of
      625 nm

Reagents
Amine-sulfuric acid stock solution:  dissolve 27 g, N, N-dimethyl-
      p-phenylenediamine oxalate (also called p-aminodimethylaniline
      oxalate) in a cold mixture of 50 ml cone. H2SOi and 20  ml
      distilled water.  Cool and dilute to 100 ml with distilled
      water.  The amine oxalate should be fresh; an old supply may be
      oxidized and discolored to a degree that results in interfering
      colors in the test.  Store in a dark glass bottle.  When this
      stock is diluted and used in the procedure with a sulfide-free
      sample, it must yield a colorless solution.
Amine-sulfuric acid reagent:  dilute 25 ml amine- sulf uric acid stock
      solution with 975 ml 1 + 1 HaSOij.  Store in a dark glass bottle.
Ferric chloride solution:  dissolve 100 g FeCla  6H20 in Uo  ml water.
Sulf uric acid solution, HaSOi* , 1 + 1.
Diammonium hydrogen phosphate solution:   dissolve ^00 g (NHOaHPOi, in
      800 ml distilled water.
Methylene blue solution I:  use USP grade dye or one certified by the
      Biological Stain Commission.   The dye content should be reported
      on the label and should be Qh percent or more.  Dissolve 0.1 g
      in distilled water and make up to 1  .   This solution will be
      approximately the correct strength, but because of variation be-
      tween different lots of dye,  standardize against sulfide solutions
      of known strength and adjust  its concentration so that 0.05 ml
      (l drop) =1.0 mg/ sulfide.
      Standardization:  put several grams of clean, washed crystals of
      sodium sulfide, Na2S '  9H20,  into a small beaker.  Add somewhat
      less than enough water to cover crystals.  Stir occasionally for
      a few minutes, then pour the solution into another vessel.  This
      solution reacts slowly with oxygen, but the change is unimportant
      in a period of a few hours.  Make the solution daily.  To 1 
      distilled water add 1 drop of solution and mix.  Immediately
  References may be found on page 3-2^7,


                               3-238

-------
      determine the sulfide concentration by the methylene blue  procedure
      and "by the titrimetric procedure.   Repeat the procedures,  using
      more than 1 drop of Na2S solution or smaller  volumes of water, un-
      til at least five tests have "been made, with  a range of sulfide
      concentrations between 1 and 8 mg/.  Calculate the average
      percent error of the methylene blue result as compared to  the
      titrimetric result.  If the average error is  negative, that  is,
      the methylene blue results are lower than the titrimetric  results,
      dilute the methylene blue solution by the same percentage, so that
      a greater volume will be used in matching colors.   If the  methylene
      blue results are high, increase the strength  of the solution by
      adding more dye.
Methylene blue solution II:  Dilute 10.00 ml of the adjusted methylene
      blue solution I to 100 ml.
Sodium hydroxide solution, NaOH, 6 !J.
Aluminum chloride solution, 6 II:  because of the hygroscopic and caking
      tendencies of this chemical, purchase 100-g (or lA-lb) bottles
      of the hexahydrate, Aids '  6H20.   Dissolve the contents of  a
      previously unopened 100-g bottle in lUU ml distilled water (or
      the contents of a lA-lb bottle in l6U ml distilled water).
Sample preparation
            Filtratipn is the routine method to distinguish between

soluble and particulate phases for most chemicals in water.   However,
this method is not desirable for sulfide because of its  labile nature.
It is possible that sulfides could be oxidized or volatized during the
filtration process.  Therefore, the following flocculation procedure
is presented to isolate soluble sulfide in the sample:
            Add 0.2 ml (k drops) 6 N_ NaOH to a 100-ml glass bottle.
            Fill the bottle with sample and add 0.2 ml (k drops)
            6 N AlCls.  Carefully stopper the bottle to  exclude  air
            bubbles.  Vigorously shake the bottle to mix the sample.
            Allow the floe to settle and draw off the clean super-
            natant.  Analyze the sample immediately for  dissolved
            sulfide or preserve with zinc acetate.   This  sample  is
            designated W2.
            NOTE:  The time allowed for the floe to settle should  be
                   kept to a minimum.
Procedure
            Decant as much water as possible from the preserved

sample without disturbing the precipitate.  Refill  the bottle with
distilled water (this step tends to remove interferences  that may  be
present in the original site water) and resuspend the precipitate.


                                3-239

-------
            Transfer a 7-5-ml sample from the well-mixed suspension to
each of two matched test tubes,  using a special widetip pipet  or  filling
to the marks on the test tubes.   To one tube, add 0-5 ml amino-sulfuric
acid reagent and 0.15 ml (3 drops)  FeCla solution.   Mix immediately by
slowly inverting the tube a single  time.  To the second tube,  add 0.5 ml
50 percent HaSOi* and 0.15 ml (3  drops) FeCla solution.  Mix the second
tube.  Allow 3 to 5 min for color development, then add 1.6 ml (NHi^HPOi*
solution to each test tube.  The presence of sulfide will be indicated
by a blue color in the first tube.   Allow 3 to 15 min for the  color to
stabilize (at least 10 min if a  zinc acetate preservative was  used) and
determine the absorbance of the  sample.  Zero the colorimeter  with a
portion of the sample from the second test tube (sample + sulfuric acid+
FeCls).  Determine the absorbance of the sample at  625 nm.
            The reaction between sulfide and the amine-sulfuric acid
reagent produces methylene blue, which is measured  colorimetrically.
In order to quantify the amount  of  sulfide in the samples,  prepare
appropriate dilutions of the standard methylene blue reagents  that have
been standardized against sodium sulfide.  Record the absorbance  of
these standards relative to a reagent blank.
Calculations
            Prepare calibration  curves by plotting  the measured absor-
bance of the standard methylene  blue solutions vs.  the sulfide equiva-
lent of each solution.  A straight-line relationship should be obtained
between 0.0 and 1.0 mg/&.  Compare  sample absorbances to the standard
curve to determine the sulfide concentrations.
            Report the results of Wl sample analysis as total  sulfide
and the results of W2 sample analyses as soluble sulfide.

Method 2:  Iodine Titrimetric1
Reagents
Hydrochloric acid, HC1, 6 N.
Standard iodine solution, 0.0205 N.:  dissolve 20 to 25 g potassium
      iodide, KI, in a little water and add 3.2 g iodine.  After  the
      iodine has dissolved, dilute  to 1000 ml and standardize  against
      0.0205 N sodium thiosulfate,  using starch solution as indicator.

-------
Standard sodium thiosulfate solution, 0.0205 N..
Starch solution.
Sodium hydroxide solution, NaOH, 6 N_.
Aluminum chloride solution, 6 N_:  because of the hygroscopic  and caking
      tendencies of this chemical, purchase 100-g (or lA-Ib) bottles
      of the hexahydrate, Aids   6H20.   Dissolve the contents of a
      previously unopened 100-g bottle in ikh ml distilled water (or  the
      contents of a lA-lb bottle in 1.6k ml distilled water).
Sample preparation
            The separation of soluble and particulate sulfide is
accomplished by alum flocculation.  Add 0.2 ml (k drops)  6 II  NaOH to
a 100-ml glass bottle.  Fill the bottle with sample and add 0.2 ml
(h drops) 6 E[ Aids.   Carefully stopper the bottle to exclude air
bubbles.  Vigorously shake the bottle to mix the sample.   Allow the
floe to settle and draw off the clean supernatant.  Analyze the sample
immediately for dissolved sulfide or preserve with zinc acetate.  This
sample is designated W2.
Procedure
            If the sulfide was precipitated with zinc and the water
decanted, conduct the titration in the original  sample bottle.  If an
unpreserved water sample is to be analyzed for sulfide, a separate
flask will be required.
            Measure from a buret a known volume  of iodine solution
that is stoichiometrically in excess of the amount of sulfide present.
Add this solution to the original sample bottle  or a 500-ml flask,
whichever is applicable.  Add distilled water, if necessary,  to bring
the volume to approximately 20 ml.  Add 2 ml 6 N_ HC1.  If the titration
is to be performed in the sample bottle, the sample is now ready.  If
the titration is to be performed in a 500-ml flask, pipet 200 ml of
unpreserved sample into the flask, taking care to discharge the sample
below the surface of the acidified solution.
            If the iodine color disappears, add  more iodine so the
color persists.  Back titrate with sodium thiosulfate solution.  When
the solution is a pale yellow, add a few drops of starch  solution and
continue the titration until the blue color disappears.
                                3-2U1

-------
Calculations
            The stoichiometry of the reaction "between iodine and sulfide
is such that 1 ml 0.025 II iodine solution will quantitatively react with
Q.k mg of sulfide.  Therefore:
                          mg s/ = ^)0(ae- b)

where
     a = ml 0.025 N. iodine solution added
     b = ml 0.025 N thiosulfate solution used
     c = initial volume of water sample, ml

-------
                  Procedure for Sediment Samples (SID)

                                                     2
Method 1:  Distillation, Methylene Blue, Color imetric

Apparatus
Distillation apparatus, all glass.  For large samples, a suitable
      assembly consists of a 1-& pyrex distilling flask with Graham
      condenser as used for the analysis of phenols.  A section of glass
      tubing should be connected to the tip of the condenser so that it
      reaches the bottom of the collection tube

Distillate collection tubes, short-form Nessler tubes, graduated at 50
      and 100 ml

Spectrophotometer , for use at 650 my and providing a light path of 1 in.
      or greater

Reagents

Nitrogen, water-pumped.
Zinc acetate, 2 N:  dissolve 220 g of Zn(C2H302)2  2H20 in distilled
      water and dilute to 1 & .

Zinc acetate, 0.2 N_:  add several drops of acetic acid to 100 ml of 2 N_
      zinc acetate solution and dilute to 1 A .
Sulfuric acid solution, 1:1:  add, cautiously, 500 ml of cone.
      to 500 ml of distilled water in a 1-& flask.  Mix continuously
      and cool under running water while combining reagents.  Cool
      solution before using.

Dilute sulfuric acid solution, approximately 0.1 N_:  dilute 5 ml of
      1:1 H2SOit to 1 H with distilled water.
Stock amine solution:  dissolve 2.7 g of N, W-dimethyl-p-phenylenediamine
      sulfate and dilute to 100 ml with 1:1 H2SOn solution.  This
      solution is stable for approximately 1 week.

Working amine solution:  dilute 2 ml of stock amine solution to 100 ml
      with 1:1 H2SOi* solution.  Prepare fresh daily.
Ferric chloride solution:  dissolve 100 g of FeCla ' 6H20 in hot dis-
      tilled water and dilute to 100 ml.  Cool before use.

Standard potassium biniodate solution, 0.025 N_:  accurately weigh
      out 0.812*1 g 101(103)2 and dissolve in distilled water.  Dilute
      to 1 I.

Standard sodium thiosulfate titrant, 0.025 N_:  dissolve 6.205 g Na2S20s 
      5H20 in distilled water and dilute to 1 &.  Preserve with 5 ml
      chloroform.  Standardize against standard potassium biniodate
      using starch as an indicator.

Potassium iodide solution:   dissolve 5 g of KI in distilled water and
      dilute to 100 ml.


                                3-2U3

-------
Treated hydrochloric acid:   place one or two strips of aluminum in a
      small beaker of cone.  HC1.   After violent reaction,  the acid is
      poured off and is ready to  use.
Oxygen-free dilution water:   pass nitrogen gas through a sufficient
      quantity of distilled water for dilution requirements.   A minimum
      of 10 min is required to displace oxygen in the water.
Sodium sulfide, reagent, crystal.
Procedure
            Prepare 0.01 IT sulfide solution as follows: weigh out
approximately 1.2 g of large crystal Na2S   9 H20.  Wash the  crystals
several times with distilled water.   Discard the washings  and add the
washed crystals to 975 nil of nitrogen-saturated distilled  water.  Dilute
to 1 &.  The exact concentration  of this stock solution is determined
by reacting the sulfide with an excess of iodine to give free sulfur
and titrating the unreacted iodine with sodium thiosulfate.
            Pipet 20 ml of stock  sulfide solution into 100 ml of
oxygen-free water.  Add 5 ml of KI solution, 20 ml of 0.025 N KH,(103)2
solution, and 10 ml of 0.1 N H2SOit.   Titrate with 0.025 N  Na2S203
solution using starch as an endpoint indicator.  Carry a blank through
the procedure and calculate the amount of reacted iodine from the
difference between the blank and  standard titrations.  Since  1 ml of
0.025 N KH(I03)2 is equivalent to 0.^00 mg of sulfide ion, calculate
the sulfide concentration in the  stock solution.  Calculate the volume
of stock solution that contains 0.2 mg sulfide and add this amount to
900 ml of oxygen-free water.  Dilute to 1 H.  This is the  working
standard containing 2 yg S/ml.
NOTE:  Sulfide solutions are extremely unstable and must be prepared
       fresh and used immediately.  Stability is increased by using
       nitrogen-saturated water for dilution.
            Prepare a standard curve by dilution of the working
sulfide solution.  Pipet 20 ml 0.2 N_ Zn(C2H302)2 into a series of 50-ml
Nessler tubes.  Add the required  amounts of sulfide solution  to each
Hessler tube, taking care to pipet the solution below the  Zn(C2H302)2
level.  Dilute to 50 ml with oxygen-free water.
            Equilibrate the temperature of the standards to 23 to 25C
using a water bath while the colorimetric reagents are added.  Add 2 ml

                                3-2UU

-------
dilute amine-sulfuric acid solution to the standard,  mix,  and add
0.25 ml (5 drops) FeCls solution.   Mix the solution and allow 10 min
for color development.  Measure the absorbance at 650 nm.
            To process samples, set up the distillation apparatus.
The transfer tube from the condenser should reach to  the bottom of
the distiHated collection tube.  The condenser should be  attached  in
such a manner that it can be easily moved up or down  when  diluting  the
distillate or adding reagents.
            Pipet 20 ml of 0.2 NZn(C2H302)2 into a 100-ml Nessler
tube and lower the condenser so that the transfer tubing reaches below
the level of the liquid.  Attach a distilling flask and pass  nitrogen
gas through the system for at least 10 min.
            Add an aliquot of field moist sediment sample  (SID)  to  the
distillation flask.   The sample should not contain more than  50 JJg  of
sulfide.  Bubble nitrogen gas through the sample to remove any oxygen
dissolved in the sample.  A small amount of sulfide may be driven over
by the gas, so be sure that the only exit is through  the zinc acetate
solution in the collecting tube.
            Discontinue nitrogen evolution and add rapidly several
boiling stones, 2 drops of methyl orange indicator, and enough treated
HC1 to change the color from orange to red.   Stopper  as quickly as
possible and heat slowly.  The slower the heating rate, the greater the
contact time between the evolved H2S and Zn(CaH302)2  and the  less chance
of sulfide loss.  Distill the solution until approximately 20 ml of
distillate has been collected (roughly 5 to 8 min after the solution
commences to boil).   Turn off heat and remove the stopper  in  the
distillation flask to keep the distillate from being  sucked back up the
condenser.  Raise the transfer tube above the 50-ml mark on the collec-
tion container and dilute the solution to 50 ml.
            Place the distillates in a water bath at  23 to 25C.  Add
2 ml dilute amine solution and mix.  Add 0.25 ml (5 drops) FeCla solu-
tion and mix.  Allow 10 min for color development and measure sample
absorbance at 650 nm.
                                 3-2U5

-------
Calculations
            Prepare a standard curve by plotting absorbance  of  the
standards vs.  sulfide concentration.   Determine the sulfide  concentra-
tion of the sample distillate by comparing sample absorbance with the
standard curve.   Calculate the sulfide concentration of  the  sediment
sample as follows:
                    ah   t  +     ^\   (C)(0.05)(1000)
                 mg S/kg (wet weight) = r^~	-
                    Oh   I A       T.4. V   (C)(0.05)(1000)
                 mg S/kg (dry weight) =    (g)($ s)	
where
        C = sulfide concentration in distillate, mg/
     0.05 = sample volume of distillate, & (as written)
        g = wet weight of sediment aliquot, g
      % S = percent solids of sediment as a decimal fraction
                                3-2^6

-------
                              References
1.  American Public Health Association.  Standard Methods for the Exami-
    nation of Water and Wastewater_. l^th Edition.  APHA; New York, New-
    York.  1193 p.  (19T6).
2.  Great Lakes Region Committee on Analytical Methods.  "Chemistry
    Laboratory Manual for Bottom Sediments."  U. S. Department of the
    Interior, Great Lakes Region; Chicago, Illinois. 96 p. (1968).
                                3-21*7

-------
ORGANIC ANALYSIS

     Carbamates
     Chlorinated Phenoxy Acid Herbicides
     Oil and Grease
     Chlorinated Hydrocarbons
          BHC
          Lindane
          Heptachlor
          Aldrin
          Heptachlor Epoxide
          DDE
          Dieldrin
          Endrin
          DDT
          Chlorobenzilate
          PCB
          Malathion
          Diazinon
          Parathion
      Organophosphorous Pesticides
      Polynuclear Aromatic Hydrocarbons
      Phenolics
                                3-21*8

-------
                              CARBAMATES
                    (N-methylcarbamate Pesticides)

            Carbamates have received increased usage because of the
concern over the persistence of chlorinated hydrocarbons.   Carbamates
are more acutely toxic than chlorinated hydrocarbons but also degrade
             i.y,
more rapidly.    These compounds generally attack the nervous system
by deactivating the enzyme chloinesterase.  A list of N-methylcarbamates,
their chemical name, and producers is presented in Table 3-19.
            Two methods are available for the quantification of carba-
mates.  One method2 involves extraction with methylene chloride,
preparation of pentafluorobenzyl bromide derivatives, and quantification
using a gas chromatograph with an electron capture detector.  The second
method utilizes the enzyme deactivation property of carbamates.   Samples
are extracted and exposed to a cholinesterase substrate (3,3-dimethyl-
butyl acetate).  The change in substrate activity is inversely propor-
tional to the carbamate concentration of the sample.  This method is
nonspecific and noncarbamate compounds that inhibit enzyme activity will
produce a positive interference.

                     Sample Collection and Storage

            Samples should be collected in glass containers.  The
preferred method of sample treatment would be to extract immediately
in the field.  If this is not possible, samples should be shielded from
exposure to sunlight and stored at UC.  Samples should be extracted
as soon as possible.  A suggested flowchart for sample handling is
presented in Figure 3-37.

              Procedures for Water Samples (Wl, W2, S1A)1

Method 1:  Methylene Chloride Extraction


* References can be found on page 3-259-
                                 3-2U9

-------
                                                Table 3-19
                              Some N-methylcarbamates and Related Compounds2
ro
^ji
o
   Trade Name
   Seviri
        R
   Mesurol'
          R
   Baygon'
         R
   Landrin
          R
   Bux
      R
        R
   Mob am
             F
   Carbofuran
   Temik
        ~
   Promecarb
            R
   Banol'
        R
   Carzol
         R
   Methomyl'
           R
   Elocron'
          R
   Dimetari
          R
   Butacarb
           R
           Systematic Chemical Name*
l-Fapthalenyl-N_-methylcarbamate
3,5 Dimethyl-U- (methylthio) -phenyl-IJ-methyl-
 carbamate
2-(1-Methylethoxy)-phenyl-N-methylcarbamat e
2,3,5-Trimethylphenyl-N-methylcarbamate
3-(l-Ethylpropyl) phenyl-N-methylcarbamate and
 3-(1-Methylbutyl) phenyl-N-methylcarbamate
Benzo[b]thiophene-U-yl-N-methylcarbamate
2,3-Dihydro-2,2-dimethyl-T-benzofuranyl-N-
 methylcarbamate
2-Methyl-2-(methylthio) propanol-0-((methylamino)
 carbonyl) oxime
3-Methyl-5-(1-methyllethyl)-N-methylcarbamate
2-Chloro-i|, 5 dimethylphenyl-N-methylcarbamate
N^,N-Dimethyl-N'- (3- (((methylamino-) carbonyl)
 oxy) phenyl)-methanimidamide monohydrochloride
N_-( ((Methylamino)carbonyl) 0-oxy)-ethanimido-
 thioic acid-IT-methylester
2- (1,3-Dioxolan-2-yl) -phenyl-N_-methylcarbamat e
5,5-Dimethyl-3-oxo-l-cyclohexen-l-yl-N-dimethyl-
 carbamate
3.5-Bi s(1,1-dimethylethyl)phenyl-N-methylcarbamat e
      Trade Source
Union Carbide

Chemagro Corp.
Chemagro Corp.
Shell Chemical Co.

Chevron Chemical Co.
Mobil Chemical

Niagara Chemical Co.

Union Carbide
Schering
Upjohn Co.

NOR-AM Agriculture Products

DuPont
CIBA-Geigy Chemical Corp.

CIBA-Geigy Chemical Corp.
Niagara Chemical Co.     	
   * Following Chemical" Abstracts.

-------
                Figure 3-37.      Handling  and storage of  samples for  carbamate  analysis

WATER SAMPLE
f
i
ii<
FILTER
1
STORE STORE
i 1
EXTRACT EXTRACT
1 i
ANALYZE ANALYZE
(Wl ) (W2)
(jO
^3 SAMPLE DESIGNATION Wl
PURPOSE Total WE
Cone.

^^^^
) TREATMENT
(W3)

^T"


1
EXTRACT
i
ANALYZE
(SIA)

W2 W3


DREDGE SAMPLE
^
^^
STORE WET
i

CORE SAMPLE
1



ter Soluble  Used In
Water Elutriate
Cone.
SIA
Mobile
Cone.


\

*
CORE SECTION |
*
t
FREEZE


STORE
1
EXTRACT EXTRACT
i 1
ANALYZE ANALYZE
(SID) (S3)

SID S3
Total Total
Sediment Sediment
Cone. Cone.



CONTAINER
SAMPLE TREATMENT
                             None
                                         Filter
                                                     None
                                                                  None
                                                                                                       None
                                                                                                                             Freeze
PRESERVATIVE
                                          k C
                           * C                                   i, C
                            (Minimize Air Contact.  Keep Field Moist.)
                                                                                                                             None
STORAGE TIME
            Not known with certainty.  Process as soon as possible.
DIGESTION SOLUTION
                            Methylene
                            Chloride
Methylene
 Chloride
Hethylene
 Chloride
Methylene
 Chloride
Methylene
 Chloride
SAMPLE VOLUME OR WEIGHT
                          0.1-1.0 liter   0.1-1.0 liter
                        0.1-1.0  liter
                                                                                                    10 -  50 g
                                                                                                                            10 - 50 g

-------
Apparatus

Water bath

Oxford pipettors

Centrifuge tubes

Vortex mixer
Gas chromatograph (GC), fid detector:  column oven 215C; detector 265C;
      inlet 2k5C; air 300 ml/min; hydrogen 20 ml/min; nitrogen Uo ml/
      min

GC column:  a glass U-tube, 6 mm by 2 m (lA in. by 6 ft) , packed with
      Chromsorb 101 (Johns Manville), mesh 80-100.   Condition overnight
      at 250C
Reagents

Stock buffer solution:  dissolve kk.f3 g KC1, k,I2 g sodium barbital,
      and 0.55 g foHPOi* in 200 ml water.
Working buffer:  add 20 ml stock buffer to 75 ml distilled water, adjust
      the pH to 8.0 with 0.1 N HC1, and dilute to 100 ml with distilled
      water.
Tween 20 (g) emulsifier.

3,3-dimethylbutyl acetate (DMBA):  to remove butanol impurities from
      DMBA, mix 5 parts DMBA with 1 part acetic anhydride.  Keep the
      mixture at 37C for 2k hr and wash once with water to remove the
      acetic anhydride and impurities.  Prepare a 0.2 M emulsion con-
      taining 0.2 percent emulsifier by diluting 0.72 g DMBA with 20 ml
      working buffer and adding 50 mg Tween 20 .  Adjust the emulsion
      to a pH of 8 with NaOH and dilute to 25 ml with working buffer.
      Shake well before using.  Store in refrigerator.
True cholinesterase, Type 1:   Keep refrigerated.  Add 3 ml working
      buffer to 50 um units of enzyme.

Formic acid solution:   dilute cone. (88 percent) HCHO with an equal
      volume of distilled water.
Bromine water:  dilute 0.2 ml bromine with 100 ml distilled water.
Acetone, pesticide quality:  redistill from glass.
Methylene chloride, pesticide quality:  Wash with water, dry over
      anhydrous CaCla, and redistill from glass.
Carbon disulfide, Spectro AR Grade.
Stock parathion solution:  dissolve 100 mg parathion in acetone  and  dilute
      to 100 ml.  1.00 ml = 1.00 mg.  Store in refrigerator.
Standard parathion solutions:   dilute the stock parathion solution with
      distilled water  to prepare standards containing 25, 50> and 100 ug/ml
      Store  in refrigerator.
                                3-252

-------
Procedure
            Place 1.0 ml Wl, W2, or S1A sample in a 15-ml centrifuge tube
and add 1 ml methylene chloride.  Shake the sample on a vortex mixer for
5 sec.  Add 0.1 ml bromine water and shake an additional 5 sec.  Remove
the upper aqueous layer with a pipet and discard.
            Transfer 10 yl of the organic solvent layer to a second
centrifuge tube as follows:
            Place the tip of the pipettor below the liquid surface.
            Depress the plunger to the first stop and release 6 to 8
            times to a.llow equilisatiori of the partial pressure in the
            top so that the liquid will remain until expelled by fully
            depressing the plunger.
            Evaporate the sample to dryness at 37C.  This is necessary
to allow the carbamates to dissolve in the enzyme solution.
            Add 100 yl enzyme solution.  Swirl and place in a 37C water
bath for 1 hr.
            Transfer 10 yl of sample to a clean centrifuge tube con-
taining 1 ml pH 8 working buffer.  Warm the tube to 37C and add 0.2 ml
DMBA emulsion.  Incubate the tube at 37C for 30 min.
            Stop the reaction by adding 0.1 ml formic acid solution and
2 ml carbon disulfide.  Stopper the tube and shake for 10 sec.   After
the layers separate, remove the aqueous layer with a pipet and discard.
            Inject 5 yl of the carbon disulfide layer into the gas
chromatograph for determination of 3,3-dimethylbutanol (DMB) which will
be eluted from the GC in approximately 2-1/2 min.  The DMBA peak is
eluted in approximately h min.
NOTE;  The procedure depends on the conversion of DMBA to DMB.   This is
       an enzymatic process that depends on the concentration of the
       enzyme, the concentration of the substrate, temperature, and
       pH.  Therefore, careful attention to procedural detail is
       necessary for reproducibility.
            Prepare a standard parathion curve by plotting parathion
concentration vs. percent inhibition.  Also, controls and blanks con-
sisting of all reagents except the enzyme should be analyzed to deter-
mine whether nonenzymatic DMBA conversion is occurring.
                                 3-253

-------
Calculations
            Determine the height of the DMB peak and calculate the percent
inhiMtlon caused by the sample as  follows:
                   % Inhibition = (a - c) - (b - c)
                                       (a - c)
where
     a = height of control DMB peak
     b = height of sample or standard DMB peak
     c = height of blank DMB peak
            Plot parathion concentration vs. percent inhibition on
semilog paper and report the sample concentration based on the observed
inhibition.
            This procedure is considered tentative by Standard Methods.
The results should be considered nonspecific as any compound that will
hydrolyze DMBA to DMB is indirectly detected and all compounds are
reported as an equivalerit weight of the standard carbamate used.
                                 3-251*

-------
                Procedure for Sediment Samples (SID)2'3

Method 1:  Methylene Chloride Extraction
Apparatus
Gas chromatograph equipped with an electron capture detector
GC columns
      a.  3.6 percent (W/W) OV-101 + 5 percent (W/W) OV-210 on 80-100
          mesh Chromosorb W, acid washed, DMCS treated
      b.  3 percent (W/W) OV-225 on 80-100 mesh Chromosorb W (HP)
Reagents
Prepare all reagent solutions in carefully cleaned glassware.   Do  not
      use any plastic ware in the preparation of reagents or the
      processing of samples.
Pentafluorobenzyl bromide (PFB) 1 percent (V/V) in acetone:  store in
      dark container.  Prepare every 2 weeks.
Methanolic potassium hydroxide - 10 percent (W/W):  dissolve 10 g  reagent
      grade potassium hydroxide in 100 ml pesticide grade methanol in
      dark bottle.
Potassium carbonate solution:  (a) (0.1 M_) 13.8 g K2C03  in 1 Jl deionized
      distilled water; (b) 5 percent solution 10 g KaCOa in 200 ml
      deionized distilled water.
50 percent sulphuric acid solution:  extract two to three times with equal
      volumes of benzene.
Silica gel:  deactivate by adding deionized distilled  water, 1.5 percent
      (W/W).  Store in tightly capped container.  Coburn et al.3
      specify grade 950 silica gel from Davison Chemical; Baltimore,
      Maryland 21226.
Anhydrous sodium sulphate.
Methylene chloride.
Benzene.
Isooctane (.2,2,U-trimethylpentane) .
5 percent Benzene-hexane (l:19).
25 percent Benzene-hexane (1:3).
75 percent Benzene-hexane (3:l).
Hexane.
Procedure
            Weigh. 30 g wet sediment.  Extract sample with 1000 ml
acidified ammonium acetate for 1 hr at 60C.  Place 1/2 of ammonium
acetate extract (.equivalent to 10 g sediment) in a 2-i separatory  funnel.
                                3-255

-------
Add sufficient 50 percent sulfuric acid to lower  the extract  pH. to  3
to U.  Add 10 g sodium sulfate.
            Extract the acidified solution twice  with 150 ml  methylene
chloride.  Shake the sample thoroughly for 10 min during each extraction.
Combine the extracts and wash with 75 to 100 ml 0.1 M potassium carbo-
nate for 5 min or less.
NOTE:  The potassium carbonate may be retained for  the analysis  of
       phenols and other acidic compounds,  if desired.
            Pass the methylene chloride extract through a narrow
anhydrous sodium sulfate column containing  20 g of  the desiccant.
Collect the sample in a round -bottomed flask.   Rinse the column  with
25 ml methylene chloride and add the rinse  to the sample flask.
Reduce the sample volume to approximately 5 nil using a rotary evaporator
and a hOC water bath.
            Add 2 ml 10 percent methanolic  potassium hydroxide to  the
methylene chloride concentrate.  Allow the  sample to hydrolyze overnight
at room temperature.
            Transfer the solution to a 500-ml separatory funnel  using
50 ml deionized distilled water.  Add 50 ml methylene chloride.  Shake
briefly and discard the methylene chloride  layer.
            Acidify the sample to pH < 2 with 50 percent sulfuric  acid
(approximately 0.3 to 0.5 ml).  Extract the acidified sample extract
with two 50-ml portions of pesticide grade  benzene.   Collect and combine
the benzene extracts.
            Prepare a narrow drying column  containing 10 g anhydrous
sodium sulfate.  Apply the benzene extract  to the column and collect in
a round-bottomed flask.  Evaporate the sample to approximately 1 ml using
a rotary evaporator and a 1*0 C water bath.
            Transfer the sample _ to a 15-mT~ graduated centrifuge  tube.
Rinse the round-bottomed flask with acetone and add to the centrifuge tube.
            Add 20 yl 5 percent potassium carbonate and 100 yl 1 percent
PFB reagent to the centrifuge tube.  Stopper and shake thoroughly  for
at least 3 hr.     .
            Add 2 ml isooctane to the derivatized sample and place in a
35 to kOC water bath.  Evaporate to approximately 1 ml by passing dry
                                 3-256

-------
nitrogen gas over the sample.  Add a second 2-ml portion of isooctane
and again reduce the volume to 1 ml.
            Prepare a silica gel column with 1.5 percent water:silica
gel in a disposable pipet.  Place 5 g anhydrous sodium sulfate on top
of the column.
            Place the isooctane solution on the column.   Rinse the
centrifuge tube with 1 ml hexane and add to the column.
            Elute the column with 5 nil 5 percent benzene:hexane.
Discard the eluate.
            Elute the column with 6 ml 25 percent benzene:hexane  into a
centrifuge tube.  Analyze this fraction for Metmercapturon, Carboxyl,
and Mobam using GC column a_.
            Elute the column with 8 ml 75 percent benzene:hexane  into
a second centrifuge tube.  Analyze the fraction for Propoxin,  Carbo-
furan, and Metmercapturon using GC column b_.
            Elute the column with 10 ml 100 percent benzene into  a
third centrifuge tube.  Analyze for 3-Ketocarbofuran using GC  column b_.
            Prepare standards with carbamates of interest.
            Place mixed standards in a round-bottomed flask, add  5 ml
methylene chloride and 2 ml 10 percent potassium hydroxide, and process
as a sample from the hydrolysis step.
NOTE:  Although the initial identification is based on GC analysis, it
       may be necessary to rely on mass spectrophotometery to  identify
       the gas chromatographic peaks.
Calculations
            Calculate th.e sediment concentration as follows:
                   M yg/kg (wet weight)  = E x  F x G
                M yg/kg (dry weight) =
where
A x B x C
E x F x G
Ax B x C
       A = weight in picograms of standard
       B = peak height (or area) of sample
       C = volume of sample extract, ml
       E = peak height (or area) of standard
       F = volume of extract required to produce B,  yl

                                 3-257

-------
g = wet weight of sediment initially extracted, g
S = percent solids in sediment sample (expressed as a decimal
    fraction)
M = concentration of methylcarbamate
                         3-258

-------
                              References
1.  American Futile Health Association.   Standard Methods  for  the
    Examination of Water and Waste-water.  lUth Edition.  APHA;  New  York,
    New York.   1193 p.  (1976).
2.  Walton, A.   "Ocean Dumping  Report  1.  Methods  for  Sampling  Analysis
    of Marine Sediments and Dredged Materials."   Department  of Fisheries
    and the Environment; Ottawa,  Ontario, Canada.   7^ p.  (1978).

3.  Coburn, J.  A., Ripley, B. D., and  Chan,  A. S.  Y.  "Analysis of
    Pesticide Residues by Chemical Derivatization II, n-methyl carba-
    mates in Natural Waters and Soils."   Official Analytical Chemists
    Journal 59:188-196 (1976).
                                3-259

-------
                  CHLORINATED PHENOXY ACID HERBICIDES

            Chlorophenoxyacetic acids such as 2,^-dichlorophenoxyacetic
acid C2,^-D), 2,lt,5-trichlorophenoxyacetic acid (2,1*,5-T), and silvex
[ 2-(.2,^,5-trichlorophenoxy) propionic acid] are herbicides used for veed
        1 4f
control.    Each compound may exist as a free acid or an ester.  In
addition, the ester forms may hydrolize in aquatic environments.2
            The analytical procedure consists of three steps.1'3
Residues are extracted into an organic solvent and esterified using
BF3 .  The methyl esters are then extracted into "benzene and quantified
using gas chromatography.

                      Sample Handling and Storage

            Water samples should be collected in an all-glass system.
The sample should be acidified with H2S04 to pH < 2 immediately after
collection and stored at kC in the dark.  Extraction of the samples
should begin within 12 hr of collection as the degradation of 2,U-D is
rapid in aqueous systems.3
            Sediment samples should be stored in glass or plastic  con-
tainers.  Immediate extraction of samples is recommended to minimize
the effects of sample degradation.   However, when necessary, sample
freezing at -20C has been shown to prolong the stability of 2,^-D.1*
            All sample containers should preferably be sealed with
Teflon-lined screw caps.1  An alternate method would be to use pre-
cleaned, heavy-duty aluminum foil to prevent the sample from coming in
contact with plastic caps and associated glue lining.  The aluminum
foil may be cleaned by washing in acetone, followed by rinsing with
pesticide grade hexane.3
            A flowchart for the processing of sediment and water samples
to be analyzed for chlorinated phenoxy acid residues is presented in
Figure 3-38.
* References for this section are found on page 3-277.
                                 3-260

-------
     Figure 3-38.      Handling and  storage  of  samples  for  chlorophenoxy  acetic acid analysis





, ,
AC 1 D 1 FY
1
i
STORE
1
EXTRACT
J
. . ANALYZE
V (Wl)
ro ' 	 '
SAMPLE DESIGNATIO
PURPOSE
WATER SAMPLE
f
1
FILTER
1
1
ACIDIFY
I
1
STORE
|
EXTRACT
I
ANALYZE

-------
             Procedures for Water Samples (.Wl,  2, S1A)1 '3


            Two extraction procedures are presented.   The only  differ-
ences in the procedures are the organic  solvents  used  in the  extraction
step and the operating conditions of the gas  chromatograph.   The
first procedure is a chloroform extraction and is used  by Environment
Canada.   The second procedur
Edition of Standard Methods. l
Canada.   The second procedure is listed as tentative in the
Method 1:  Chloroform Extraction3

Apparatus

All glassware must be washed in chromic acid, rinsed in dilute hydro-
      chloric acid followed by distilled water,  and then rinsed in
      acetone and hexane.  Heat treatment is carried out at  300 C  on
      all glassware except volumetric flasks and pipets.  Care must  be
      taken to ensure that the glassware is not  alkaline. Considerable
      loss at low levels of herbicide can be attributed to the alka-
      linity of the glassware
Gas chromatograph such as a Varian 2800, Microtec 220,  or equivalent.
      It should be equipped with an electron capture detector, a glass-
      lined injection port, and a recorder.  Recommended operating
      conditions are:  column temperature, 195C; injection  temperature,
      250C; detector temperature, 275C; attenuation,  16; and carrier
      flow rate, 60 ml Nz/min

Chromatographic column:  glass, 1.8 m by h mm I.D.   One of the following
      four mixtures can be used as column packing to separate and  quan-
      tify chlorinated phenoxy acid herbicides:

      a_.  McKinley and McGully's column .(196^):   k percent SE-30 and
          6 percent QF-1 on 100-120 mesh size, chromosorb W, acid
          washed and DMCS treated

      b_.  3 percent Dexil 300 GC on chromosorb W, acid  washed and  DMCS
          treated, 100-120 mesh size

      .  3 percent OV-1 on chromosorb W, acid washed and DMCS treated,
          100-120 mesh size
      d..  Chau-Wilkinson Column (1972):  U percent OV-101/6  percent  OV-210
          on chromosorb , acid washed, HDMS treated, 80-100 mesh  size

Pipets:  Pasteur, disposable, 1^0 mm long by 5 mm I.D., glass

Graduated centrifuge tubes:  15 ml with ground glass stoppers
Flasks:  volumetric, 1.0, 2.0, 10, and 100 ml

Flasks:  round bottomed, 300 ml

Evaporator, rotary


                                3-262

-------
Micro-syringes:  Hamilton, 10 yl for injections
Oven (.capable of maintaining 300C)
Separatory funnels:  2-& and 500-ml sizes with. TFE-fluorocarbon stop-
      cocks and taper ground glass stoppers, Kontes, or equivalent
Reagents
Check all reagents for purity by the gas chromatograph procedure.
      Much time and effort is saved by selecting high-quality reagents
      that do not require further preparation.  Some purification of
      reagents may be necessary as outlined below.   If more rigorous
      treatment is indicated, obtain the reagent from an alternate
      source.
Benzene:  pesticide quality, distilled in glass.
Sodium sulfate, anhydrous, granular:  store at 130 C.
Sodium sulfate solution:  dissolve 50 ml anhydrous  Na2SOit  in distilled
      water and dilute to 1 H.
Sodium sulfate, acidified:  add 0.1 ml cone. H2S04  to 100 g Na2SOit slur
      ried with enough ethyl ether to just cover the solid.  Remove the
      ether by vacuum drying.  Mix 1 g of the resulting solid with 5 ml
      distilled water and confirm that the mixture  has a pH value below
      U.  Store at 130 C.
Sulfuric acid:  HaSOi*, cone.
Boron trifluoride-methanol:  lU percent boron trifluoride by weight.

Florisil adsorbent:  60-100 mesh, factory activated at 650 C.  The
      florisil is heated to 130 C for 1 hr and stored in a desiccator
      prior to use.  Each batch is checked for activity and for con-
      tamination.
Glass wool:  filtering grade, acid washed.

Analytical standards:  MCPA; MCPA methyl ester; 2,^,5,-T; 2,U-D; 2,U,5-T
      methyl ester; 2,U-D methyl ester; silvex, silvex methyl ester;  all
      at least 98+ percent purity (available from Dow Chemical).
Stock herbicide solutions:  dissolve 100 g herbicide or methyl ester  in
      60 ml ethyl ether; dilute to 100 ml in a volumetric flask with
      hexane.  1.00 ml = 1.00 mg.

Intermediate herbicide solution:  dilute 1.0 ml stock solution to 100 ml
      in a volumetric flask with a mixture of equal volumes of ethyl
      ether and benzene.  1.00 ml = 10.0 yg.
Standard solution for chromatography :  prepare final concentration of
      methyl ester standards in benzene solution according to the
      detector sensitivity and linearity.

Hexane .
Chloroform.

Methanol.

                                3-263

-------
Procedure
            Acidify a 1000-ml water sample to pH 2.0 with cone.
Transfer the sample to a 2000-ml separatory funnel.
            Rinse the sample container with 50 ml chloroform and add
the rinsing to the separatory funnel.   Shake the mixture thoroughly for
a minimum of 1 min.  Allow 5 min for complete separation to occur and
draw off the bottom .chloroform layer into a clean 500-ml separatory
funnel.  Should an emulsion form during the extraction procedure, it can
usually be broken by adding small quantities of 2-propanol, acetone, or
a saturated Nad solution.
            Repeat the extraction a second and a third time using 50-ml
portions of chloroform.  Combine the extracts in the 500-ml separatory
funnel.  Wash, the chloroform extract with 100 ml glass-distilled water.
Remove the aqueous layer, making sure that it is slightly acidic.
            Dry the combined chloroform extract over acidified sodium
sulfate for 10 min.  The extract should not remain in contact with the
sodium sulfate layer for more than 1/2 hr.
            Concentrate the dried extract on a rotary evaporator to a
volume of approximately 5 ml.  Add 10 ml of methanol and evaporate again
to 5 ml.  Continue this procedure until all traces of chloroform have
been removed.  Transfer the resultant methanol solution to a 15-ml
graduated centrifuge tube and concentrate to 1 ml under a gentle stream
of nitrogen.
            Add 0.5 ml BFs-methanol complex to the sample in a graduated
centrifuge tube and heat in a water bath at 50C for 30 min.   Allow the
reaction mixture to cool to room temperature.
            Add 5 ml of 5 percent aqueous sodium sulfate solution to the
centrifuge tube.  Extract the methyl esters with two successive 2-ml
portions of hexane.  Concentrate the hexane extract  to 1 ml under a
stream of dry nitrogen.
            Prepare a small column by plugging a disposable pipet with
glass wool.  Pack the column with 2.0 cm of florisil and 2.0 cm neutral
anhydrous sodium sulfate.  Pass the hexane phase containing the methyl
esters of the phenoxy acid herbicides through the column.  Elute the
herbicides with 10 ml of benzene.
                                3-264

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            Concentrate the benzene solution to 0.5 ml under a stream of
dry nitrogen and quantitatively transfer the solution to a 1-ml volu-
metric flask.  This solution is now ready for quantification using gas
chromat ography.
            Preliminary identification is achieved via electron
capture GLC using at least two different stationary phases of different
polarity.  The identity of the herbicide is based on the retention time
relative to aldrin.
            Confirmation of residue identity may be achieved by one of
the following methods:
            a_.  Transesterification to higher molecular weight esters.5
            b_.  Thin-layer chromatography utilizing silica gel G as the
                absorbent and benzene as the mobile phase.  Development
                can be achieved using a silver nitrate spray reagent.6
            c_.  Mass spectroscopy.
Calculations
            The concentration of chlorinated phenoxy acid herbicides in
the water sample is calculated as:
                          P - A x B x C     -3
                          P ~ E x F x G   10
where
     P = concentration of chlorinated phenoxy acid herbicides,  pg/
     A = weight in picograms of standard
     B = peak height (or area) of sample
     C = volume of sample extract, ml
     E = peak height (.or area) of standard
     F = volume of extract required to produce B, yl
     G = volume of water sample initially extracted, H
Method 2:  Ethyl Ether Extraction1
Apparatus
Gas chromatograph such as a Varian 2800, Microtec 220, or equivalent.
      It should be equipped with an electron capture detector, a glass-
      lined injection port, and a recorder.  The following operating
      conditions are recommended:  injection temperature, 215C; oven
      temperature, l85C; column temperature, l85C; and a carrier gas
      flow of 70 ml/min in a 6.k mm-O.D. column
Chromatographic column:  the use of two columns is suggested for
                                 3-265

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      identification and confirmation.  One column is packed with 1.5
      percent OV-1T and 1.95 percent QJF-1 on a 100/120 mesh Gas Chrom
      Q.  The second column is packed with 5 percent OV-210 on a 100/
      120 mesh Gas Chrom Q.

Pipets:   Pasteur, disposable, 1^0 mm long by 5 mm I.D., glass

Micro-syringes:   Hamilton, 10 yl for injections
Oven (capable of maintaining 300C)

Evaporator concentrator:  Kuderna -Danish, 250-ml flask and 5 -ml volu-
      metric receiver, Kontes or equivalent
Snyder columns:   three-ball macro, one-ball micro

Separatory funnels:  2- and 60-ml sizes with TFE-fluorocarbon stop-
      cocks and taper ground glass stoppers, Kontes, or equivalent
Sand bath:  fluidized (TeCam or equivalent) or water bath
Erlenmeyer flask:  250 ml, with ground glass mouth to fit Snyder
      columns
Reagents

Check all reagents for purity by. the gas chromatograph procedure.
      Much time and effort is saved by selecting high-quality reagents
      that do not require further preparation.  Some purification of
      reagents may be necessary as outlined below.  If more rigorous
      treatment is indicated, obtain the reagent from an alternate
      source.
Ethyl ether:  reagent grade.  Redistill in glass after refluxing over
      granulated sodium-lead alloy for k hr.
Benzene:  pesticide quality, distilled in glass.
Sodium sulfate:   anhydrous, granular.  Store at 130 C.
Sodium sulfate solution:  dissolve 50 ml anhydrous NaaSOi, in distilled
      water and dilute to 1 .
Sodium sulfate, acidified:  add 0.1 ml cone. H^SOt, to 100 g
      slurried with enough ethyl ether to just cover the solid.  Remove
      the ether by vacuum drying.   Mix 1 g of the resulting solid with
      5 ml distilled water and confirm that the mixture has a pH value
      below h.  Store at 130 C.
Sulfuric acid:  HaSOit, cone.

Sulfuric acid, H2SOi,, 9 N_:  store in refrigerator.

Potassium hydroxide solution:  dissolve 37 g K.OH in distilled water and
      dilute to 100 ml.

Boron trifluoride-methanol:  lU percent boron trifluoride by weight.

Florisil adsorbent:  60-100 mesh, factory activated at 650C.  The
      florisil is heated to 130C for 1 hr and stored in a desiccator
      prior to use.  Each batch is checked for activity and for contami-
      nation.
                                3-266

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Glass wool:  filtering grade, acid washed.
Analytical standards:  MCPA; MCPA methyl ester; 2,1|,5-T; 2,h-D;
      2,1|,5-T methyl ester; 2,U-D methyl ester; silvex; silvex
      methyl ester; all at least 98+ percent purity (available
      from Dow Chemical).
Stock herbicide solution:  dissolve 100 mg herbicide or methyl ester
      in 60 ml ethyl ether; dilute to 100 ml in a volumetric flask
      with hexane.  1.00 ml = 1.00 mg.
Intermediate herbicide solution:  dilute 1.0 ml stock solution to 100 ml
      in a volumetric flask with a mixture of equal volumes of ethyl
      ether and benzene.  1.00 ml = 10.0 Ug.
Standard solution for chromatography :   prepare final concentration of
      methyl ester standards in benzene solution according to the
      detector sensitivity and linearity.
Procedure
            Measure 1  of a Wl or W2 sample using a graduated cylinder.
Acidify to pH 2 with cone. I^SOit and transfer to a 2- separatory
funnel.  Add 150 ml ethyl ether to the separatory funnel and shake
vigorously for 1 min.  Let phases separate for 10 min.  If emulsions
form, drain off the aqueous layer, invert the funnel, and shake rapidly.
NOTE:  Vent the funnel frequently to prevent excessive pressure buildup.
            Collect the extract in a 250-ml ground glass-stoppered
Erlenmeyer flask containing 2 ml KOH solution.  Repeat the extraction
with two 50-ml portions of ethyl ether.  Combine the extracts in the
Erlenmeyer flask.
            Add 15 ml distilled water and a small boiling stone to the
flask.  Attach a three-ball Snyder column.  Remove the ether on a steam
bath and continue heating for a total of 60 min.
            Transfer the concentrate to a 60-ml separatory funnel.
Extract the sample with 20 ml ethyl ether and discard the ether layer.
Repeat the ether extraction and again discard the ether layer.  The
herbicides are retained in the aqueous phase.
            Acidify with 2 ml cold (bC) 1+3 HzSOi*.  Extract once with
20 ml ethyl ether and twice with 10 ml ethyl ether.  Collect the extracts
in a 125-ml Erlenmeyer flask containing 0.5 g acidified anhydrous
Let the extract remain in contact with the NaaSOi* for at least 2 hr.
            Fit a Kuderna-Danish apparatus with a 5-nil volumetric

                                3-267

-------
receiver.  Transfer the ether extract to the Kuderna-Danish apparatus
through a funnel plugged with glass wool.  Use literal washing of ether.
Crush any hardened NaaSOi, with a glass rod.  Before concentrating, add
0.5 ml benzene.
            Reduce the volume to less than 1 ml on a sand bath or on a
steam bath heated to 60 to 70C.  Attach a Snyder microcolumn to the
Kuderna-Danish receiver and concentrate to less than 0.5 ml.
            Cool and add 0.5 ml boron trifluoride-methanol reagent.
Use the small one-ball Snyder column as an air-cooled condenser and
hold the contents of the receiver at 50C for 30 min in the sand bath.
Cool and add enough NaaSOii solution so that the benzene-water interface
is in the neck of the Kuderna-Danish volumetric receiver flask (about
^.5 ml).  Stopper the flask with a ground-glass stopper and shake
vigorously for about 1 min.  Let stand for 3 min for phase separation.
            Pipet the solvent layer from the receiver to the top of a
small column prepared by plugging a disposable Pasteur pipet with glass
wool and packing with 2.0 cm Na2SOi* over 1.5 cm florisil adsorbent.
Collect the eluate in a 2.5-ml graduated centrifuge tube.   Complete the
transfer by repeatedly rinsing the volumetric receiver with small
quantities of benzene until a final volume of 2.0 ml of eluate is
obtained.  Check calibration of centrifuge tubes to ensure that the
graduations are correct.
            Analyze the benzene extract by gas chromatography using at
least two columns.  Injections of 5 to 10 yl should be sufficient for
this purpose.
            Inject standard herbicide methyl esters frequently to ensure
optimum operating conditions.  Always inject the same volume.   Adjust
the volume of sample extract with benzene, if necessary, so that the
heights of the peaks obtained are close to those of the standards.
(.If a portion of the extract solution was concentrated, the dilution
factor  D  is less than 1; if it was diluted, the dilution factor
exceeds 1.)
            The identity of the residue may be confirmed by:
            a_.  Transesterificiation.
            b_.  Thin-layer chromatography,
                               3-268

-------
            c_.  Mass spectroscopy.

Calculations

            Compare the peak height of a standard to the peak height

of the sample to determine the amount of the herbicide injected.

            Calculate the concentration of herbicides:

                                A x B x C x D
                                  Ex F x G

where

     A = weight of herbicide standard injected
     B = peak height of sample, mm

     C = extract volume, yl

     D = dilution factor

     E = peak height of standard, mm

     F = volume injected, yl

     G = volume of sample extracted, ml

            To determine recovery efficiency,  add known amounts  of
herbicides to a 1-& water sample, carry through the same procedure as

the samples, and determine recovery efficiency.  Periodically determine

recovery efficiency and a control blank to test the procedure.   Analyze

one set of duplicates with each series of samples as a quality control

check.
NOTE 1:  Extraneous matter, especially in highly colored water samples,
         is a potential interference.  The cleanup procedure  described
         here will usually eliminate this source of interference.   Many
         organic compounds can interfere with  the analysis, however.
         Boron trifluoride-methanol reagent is used to advantage because
         it reacts specifically with carboxylic acids, whereas 'diazo-
         methane may react with phenols and other organics with  rela-
         tively active hydrogens.  All reagents must be thoroughly
         checked and any interferences from this source eliminated.
NOTE 2:  Strict attention is required of the analyst to obtain repro-
         ducible and satisfactory recovery. In the steps where  solvents
         are evaporated, extreme care must be  exercised, especially when
         working with the methyl esters.  The  extracts should never be
         taken to dryness as the esters are extremely volatile.
NOTE 3:  Care must be taken to ensure that the tubes are tightly capped
         and remain so after introduction of the BFs-methanol reagent.
         The temperature should be about 50C  for good yields.   The
         methylation is a very critical step in the procedure.
                                3-269

-------
NOTE k:  Sodium sulfate has "been questioned due to its relative reten-
         tative property for 2,^-D.  However, if that reagent is not
         basic, the recoveries are good.
                                3-270

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              Procedure for Sediment Samples (.SID, S3)7'8


Method 1:  Acetone-Hexane Extraction
Apparatus
Gas chromatograph equipped with an electron capture detector and a
      recorder.  Operating conditions are:  column temperature, 200C;
      injection port, 230C; and detector temperature, 3^0 C.  Use
      5 percent methane and 95 percent argon for both carrier gas
      flov (ho ml/min) and make-up gas flow (.20 ml/min)

Chromatograph column, glass U-Tube, 2 m by 3.5 nim O.D.
Two column packings have been shown to be useful for separating and
      quantifying chlorinated phenoxy acid herbicides:
      a_.  11 percent OV17 + QF-1 mixed phase by weight on 80/100 mesh
          Gas Chrom Q available from Applied Science
      b_.  3 percent QV1T on Chromosorb W, HP 80/100 mesh available
          from Applied Science
Ultrasonic homogenizer:  such as the Sonicator Cell Disrupter Model
      -375 with a solid disrupter form (#280-0.75")-  This is
      available from Heat Systems-Ultrasonic, Inc., 38 East Mall,
      Plainview, Long Island
Solvent evaporator:  such as the Buchi Rotovap
Centrifuge tube heater:  such as the Kontest Tube Heater block set at
      ^0C combined with a gentle stream of pure N2  gas for controlled
      evaporation
All glassware must be thoroughly washed with laboratory soap, rinsed with
      tap HaO, and rinsed with diluted HC1, followed by distilled HaO,
      acetone, and hexane.   Heat treatment is carried out at 300C on
      all glassware except volumetric flasks and disposable pipets
NOTE:  Glassware must be acidic.  Considerable loss at low levels of
       herbicide can be attributed to alkalinity of the glassware.
Oven:  capable of maintaining 300C
Pipets:  disposable
Beakers:  100 ml
Beakers:  200 ml
Tubes:  graduated centrifuge, glass with ground-glass stopper
Flasks:  flat bottomed, 500 ml
Funnels:  coarse, sintered glass, with ground-glass joints
Flasks:  suction with ground-glass joints
Funnels:  separatory, 500ml
                                3-271

-------
Funnels, powder:  glass

Column:  chromatographic (JLO mm I.D. x 300 mm) vi.th. coarse frit and
      stopcock.  Reservoir at top (.28 mm I.D. x 150 mm)

Syringe:  Hamilton, injection, 10 yl

Reagents
All solvents must be of pesticide quality and should be checked before
      use.  All chemicals must be of highest purity and should be
      suitably pretreated as required.

Benzene.

Hexane.

Methylene Chloride.

Acetone.

1:1 Acetone:Hexane

Acidified organic-free H20:  add hexane (50 ml) to distilled H20 (5 &)
      and stir for k hr on a magnetic stirrer at maximum speed.  Trans-
      fer to a large separatory funnel and remove the vater layer
      into storage bottles.  Add cone. HC1 (.2 ml/).
HC1, cone, (..analyzed reagent grade or better).

Celite filter aid.

Silica gel ignited at 650C overnight, homogenized vith 5 percent
      organic-free water for 2 hr prior to use.
Anhydrous sodium sulfate (.ASC grade or better) ignited at 650C over-
      night.
Acidic sodium sulfate:  acidify acetone (250 ml) with cone. HC1 to pH It.
      Place ignited Na2SOi, into a clean porcelain tray and homogenize
      with the acetone solution.  Allow to dry overnight in a fumehood.
      Place in storage bottles.
5 percent Na2SOi solution:  dissolve heat-treated Na2SOij (50 g) in
      organic-free H20 and dilute to 1 .
Analytical standards:  2,lt-DP; 2,It-D; 2,It,5-T; silvex and 2,It-DB acids
      and esters, all 99+ percent pure (available from Polyscience or
      as Environmental Protection Agency reference standards).
Boron trifluoride (.lit percent)-methanol complex esterification reagent
      (.available from Analabs).
Procedure

            Weigh a 25-g dry weight equivalent of a homogenized SID or

S3 sediment sample.  Transfer to a 250-ml beaker and slurry with acidi-
fied organic-free water.  The resultant mixture should be approximately

20 to 30 percent water.


                               3-272

-------
            Thoroughly mix the sediment slurry and carefully acidify
the sample with, k ml cone, hydrochloric acid.
NOTE:  Add acid slowly with mixing to prevent mechanical loss due to
       gas exp.ulsion.  Allow the mixture to sit 20 min, stirring
       occasionally.
            Add 5 ml 1:1 acetone:hexane mixture to the acidified sedi-
ment.  Place the ultrasonic homogenizer disrupter horn approximately
2 cm into the sample.  Activate the disrupter for 2 min in the pulsed
mode at 35 percent duty cycle with maximum output.  Allow the sediment
to settle.
            Prepare a slurry of 1:1 acetone/hexane and celite.  Pour
the slurry into a sintered glass funnel which is connected to a suction
flask.  Activate vacuum to remove the acetone/hexane from the celite
filter bed.  Discard the acetone/hexane.
            Decant the supernatant solvent from the sample into the
funnel and apply a vacuum to collect the extract in the suction flask.
Retain the solids for a second extraction.
            Add 75 ml 1:1 acetone:hexane to the sediment.  Mix with the
ultrasonic homogenizer, allow the sediment to settle, and filter through
the celite filter bed.
            Transfer the combined extract to a 500-ml separatory funnel.
Add 100 ml acidified organic-free water and shake for 1 min.  Release
the pressure frequently.  Allow the layers to separate and transfer the
aqueous layer back to the suction flask.
            Slowly pour the solvent layer through a glass powder funnel
plugged with, glass wool and containing approximately 2 cm of acidic
Na2SOit.  Trap the solvent in a 500-ml flat-bottomed flask.
            Return the aqueous layer from the suction flask to the
separatory funnel.  Rinse the suction flask with 75 ml methylene
chloride and add the rinses to the separatory funnel.  Shake for 1 min
and allow the layers to separate.
NOTE:  If an emulsion persists, leave it with the aqueous layer.
            Decant the lower solvent layer through the Na2SOit funnel and
into the 500-ml flat-bottomed flask.
            Extract the aqueous phase with a second 75-*nl portion of

                                3-273

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methylene chloride.  Filter the methylene chloride phase  through the
acidified NazSOi* funnel and combine with previous extracts.
NOTE:  There should not "be any fcO in the extract.
            Transfer the extract to a Buchi evaporator  and reduce the
volume to 2 to 5 ml.  Transfer the residue to a 15-ml graduated centri-
fuge tube and evaporate to 0.5 ml.
            Add 1 ml of benzene and shake.   Reduce the  volume to
0.5 ml.  Repeat the process of adding 1 ml of benzene and reducing
the volume to 0.5 ml until the extracted residue is in  benzene and
methylene chloride has been removed.
            Add 0.2 ml Ik percent boron trifluoride-methanol  esteri-
fication reagent and shake for 1 min.  Seal tightly and place the tube
in a water bath at 50C for 30 min.
            Cool to room temperature and add 5 ml 5 percent NaaSOi*
solution.  Shake for 1 min and allow the layers to separate.
            Withdraw the top layer into a clean centrifuge tube using
a Pasteur pipet.  Add 1 ml benzene and shake.  Allow the  layers to
separate and transfer the top benzene layer to a clean  centrifuge tube.
Repeat the benzene extraction a second and a third time.
            Evaporate the final benzene extract to a volume of 0.5 ml.
Add 1 ml hexane, shake, and reduce the volume to 0.5 ml.   Repeat this
process an additional two times to bring the extract into hexane.
            Prepare a cleanup column by adding preheated  silica gel
to a height of 75 mm.  Tap the column while packing. Add 12  mm of
neutral anhydrous NaaSOi^ to the top of the column.   Elute the column
with approximately 30 ml of hexane and discard the eluant.
            Carefully transfer the 0.5-ml hexane residue  to the clean-
up column.  Rinse the centrifuge tube with three 1-ml portions of hexane
and add each rinsing to the cleanup column.  Allow the  column to elute
until the hexane layer just recedes to the top of the l^SOi*.
            Add 90 ml of hexane and elute.   Discard the eluant.
            Add 100 ml benzene and elute.  Collect the  solvent in a
500-ml flat-bottomed flask.  Elute with a second 100-ml portion of
benzene and combine with the first eluant.   Reduce the  volume to approxi-
mately 5 ml on a Buchi evaporator.  Transfer to a 10-ml volumetric flask
                                3-27U

-------
and dilute to volume with benzene.
            Preliminary identification is achieved via electron capture
GLC in which at least two different stationary phases of different
polarity are employed.  The identity of the herbicide is based on the
retention time relative to aldrin.
            Confirmation can be achieved by transesterification, thin-
layer chromatography, or mass spectroscopy.
Calculations
            The concentration of chlorinated phenoxy acid herbicides
in the sediment sample can be calculated as:
                                       A  v "D v f*
                      P (wet weight) = E x F x H

                   P (dry weight) =
                                    E x F x  H x    s
where
       P = concentration of chlorinated phenoxy  acid herbicides,  yg/kg
       A = weight in picograms of standard
       B = peak, height (.or area) of sample
       C = volume of sample extract, ml
       E = peak height (or area) of standard
       F = volume of extract required to produce B,  yl
       H = wet weight of sediment initially extracted, g
     % S = percent solids in sediment sample (expressed as a decimal
           fraction)
Remarks
            Strict attention is required of the  analyst to obtain repro-
ducible and satisfactory recovery.   In the steps where solvents  are
evaporated, extreme care must be exercised,  especially when working with
the methyl esters.  The extracts should never be taken to dryness as  the
esters are extremely volatile.
            In natural sediment samples, benzene elutes brown matter
which, upon GLC injection, remains  in the glass  sleeve liner due to its
nonvolatile nature.
            Where emulsions form at the solvent/H^O  interface, the
emulsion should remain with the H20 phase.  This allows the emulsion  to
be extracted further with methylene chloride and prevents the

                                3-275

-------
in the funnel from becoming saturated with, water.
            Any amount of water in the extract could inhibit esterifi-
cation and result in depressed recoveries.
            The procedure has been shown to produce greater than 90
percent recoveries with known standards.  The presence of organic
matter in samples can reduce recoveries.
                               3-276

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                              References
1.  American Public Health Association.   Standard Methods  for  the Exami-
    nation of Water and Wastewater Including Bottom Sediments  and
    Sludges. lUth Edition.APHA;  New York,  New York.1193 p. (1976).

2.  Junk, G. A., Richard,  J. J.,  Fritz,  J. S.,  and Svec, H. J.  "Resin
    Sorption Methods for Monitoring Selected Contaminants  in Water."
    In:   Identification and Analysis of  Organic Pollutants in  Water.
    L. H. Keith (Ed.).Ann Arbor  Science Publishers;  Ann  Arbor,
    Michigan,  pp. 135-153 (.1976).
3.  Environment Canada.  "Analytical Methods Manual."   Inland  Waters
    Directorate, Water  Quality Branch; Ottawa,  Ontario, Canada (197^).
h.  Bristol, D.  "Effects  of Storage Conditions of Residues of 2,U-D and
    2,it-DCP in Potatoes."   In:  "Accuracy in Trace Analysis:  Sampling,
    Sample Handling, and Analysis."  National Bureau of Standards
    Special Publication 1*22.  pp.  737-7^5 (1976).
5.  Yip, G.  "Confirmation of Chlorophenoxy  Acid Herbicide Residues by
    Transesterification."   J. Assn. Of fie. Anal. Chem. 51*:31*3-3l* (.1971).
6.  Chau, A. S. Y.  "Analysis of Chlorinated Hydrocarbon Pesticides in
    Waters and Wastewaters. Methods in Use in Water Quality Division
    Laboratories."  Department of the Environment, Inland  Waters
    Directorate; Ottawa, Ontario,  Canada (1972).
7.  Walton, A.  "Ocean  Dumping Report 1. Methods for Sampling  and Analysis
    of Marine Sediments and Dredged Materials."  Department of Fisheries
    and Environment; Ottawa, Ontario, Canada.  7^ P- (1978).
8.  Peake, A. A., and Lesick, H.  S.  "Procedure for the Analysis of
    Phenoxy Acid Herbicides in Sediments."   Water Quality  Branch,
    Inland Waters Directorate; Calgary,  Alberta, Canada.   9 ? (-No date).
                                  3-277

-------
                            OIL AND GREASE
            The oil and grease procedure is a gross measurement of a
fraction of the organic material that may be present in water  and sedi-
             i*
ment samples.    The procedure is operationally defined and based on the
solubility of organic matter in a nonpolar solvent under acidic condi-
tions.  Therefore, specific compounds that may be included in  an oil and
grease determination are hydrocarbons, vegetable oils,  animal  fats, waxes,
                                                 1  9
soaps, greases, and related industrial compounds. '
                      Sample Handling and Storage

            The oil and grease procedure can be performed with either
water or sediment samples.  However, the test should not be run on filtered
water samples as part of the oil and grease can be lost during the fil-
tration process.3  If it is desired to estimate the dissolved oil  and
grease fraction, a separate water sample or a separate elutriate prepa-
ration should be centrifuged.  Samples should be collected and stored  in
glass containers and preserved with sulfuric acid (pH < 2).
            Sediments may lose apparent oil and grease as a result of
drying.  Therefore, it is recommended that sediments to be analyzed for
oil and grease be stored in a field moist condition at ^C.   A schematic
flowchart for oil and grease sample handling is presented in Figure 3-39-

               Procedure for Water Samples (Wl, W2, S1A)

Method 1:  Freon Extraction1'2
Apparatus
Separating funnel, 2000 ml, with Teflon stopcock
Extraction apparatus, Soxhlet
Distilling flask, 125 ml
Water bath
Infrared spectrophotometer, double beam, recording and quartz cells

* References can be found on page 3-288.
                                 3-278

-------
             Figure 3-39.     Handling and storage  of samples  for oil and grease analysis




,,
ACIDIFY
1
STORE
I
EXTRACT
1
ANALYZE
(Wl)
UJ

WATER SAMPLE CORE SAMPLE
i ' 4
t  *
CENTRIFUGE DREDGE SAMPLE CORE SECTION
I
jL &
i r f
ACIDIFY N TR(yENT STORE WET
1 	
STORE * Elutriate should be
1
centrirugea rather tnan
A ^ , * SfanHarH Fl ufr i =fo ,
EXTRACT 	 3. ELUTRIATE* B'?*?OY EXTRACT
1 1 1
ANALYZE ANALYZE ANALYZE
(W2) (S1A) (SID)


^ SAMPLE DESIGNATION
PURPOSE


CONTAINER
SAMPLE TREATMENT
PRESERVATIVE


STORAGE TIME









Wl W2 W3 S1A SIC SID
Total Water Soluble Used In Mobile Bioavail- Total
Cone. Water Elutriate Cone. ability Sediment
Cone. Cone.
G G G G G G
None Centrifuge None None None None
H2SO,, H2SOk I|0C ')C baC 4C
pH<2 pH<2
(Minimize Air Contact. Keep Field Moist.)
Iw Iw Iw Iw Iw
DIGESTION SOLUTION
                        Freon
                                   Freon
                                                        Freon
                                                                                         Freon
SAMPLE VOLUME OR WEIGHT
                          liter
                                   1 liter
                                                        1  liter
                                                                            Variable
                                                                                         20g

-------
Balance
Reagents
Either 1:1 sulfuric acid or 1:1 nitric acid:   mix equal volumes of the
      concentrated acid and distilled water.
Freon 113, b.p. U8C; l,l,2-trichloro-l,2,2-trifluoroethane:   filter
      1-gal quantities through paper into glass containers.   The solvent
      should leave no measurable residue on evaporation.  Solvent blanks
      should be run routinely as a quality control check.  Redistill  the
      solvent when necessary.
Sodium sulfate, NaaSOi*, anhydrous crystal.
Known oil reference standard:  accurately weight about 0.05  g of known
      oil directly into a 100-ml volumetric flask.  Add 80 ml Freon and
      dissolve the oil.  If, as in the case of a heavy fuel  oil, all
      the oil does not go into solution, let stand overnight.  Filter
      through a Whatman No. Ho filter paper into a second  100-ml volu-
      metric flask and dilute to volume with Freon.
Unknown oil reference standard (10 yl = 7-69 mg oil):   pipet  15.0 ml
      n-hexadecane, 15.0 ml isooctane, and 10.0 ml benzene into a 50-ml
      glass-stoppered bottle.  Assume the specific gravity of this
      mixture to be 0.769 and maintain the integrity of the mixture by
      keeping the bottle stoppered except when withdrawing aliquots.
Procedure
            Mark the liquid level on the sample container  for later
determination of sample volume.  If the sample was not acidified at the
time of collection, add 5 nil of sulfuric acid or hydrochloric acid to the
sample bottle.  Mix the sample and measure the pH to ensure that the  pH
is 2 or lower.  Add additional acid if necessary.
            Transfer approximately 1 & of unfiltered water or centrifuged
water sample into a separating funnel.  Rinse the sample container with
30 ml Freon 113 and add the solvent washings to the separating funnel.
Shake the separating funnel vigorously for 2 min and allow the layers to
separate.
            Soak a Whatman No. kQ filter in Freon 113 and  mount in a
funnel.  Drain the Freon layer from the separating funnel  through the
solvent-moistened filter paper and into a clean collection vessel.
            Rinse the original sample container with a second 30-ml
portion of Freon, add the washings to the sample in the separatory
funnel, and extract a second time.  Drain the organic  layer through the
filter paper and combine with the first extract.  Repeat the  entire
                                 3-280

-------
procedure with a third 30-ml portion of Freon.

            The final combined extract should be  a clear  solution.   If

an emulsion has formed, add approximately 1 g anhydrous sodium sulfate,

Na2SOi, to the funnel cone and refilter the combined extract.   Use  addi-

tional sodium sulfate as required.

            Rinse the tip of the separating funnel, the filter paper,

and the funnel vith 10 to 20 ml Freon.  Collect the washings and  add to

the sample extract in the collection vessel.

            The extracted material  can be quantified as oil and grease
using one of the following methods:   (a) infrared spectrophotometery or

(b) gravimetry.

            a_.  Infrared spectrophotometry .* '2  If this method of quan-
                tification is to be  used, it  would be convenient  to
                collect the sample  extracts and washings  in a  100-ml
                volumetric flask.   Dilute the combined extracts to
                volume with Freon.
                Prepare calibration standards using either the known oil
                reference standard  or the unknown oil reference standard.
                (.A known oil is defined as the only grease and/or oil
                component in the samples being analyzed.  An unknown oil
                is defined as the grease and/or oil component(s)  in the
                sample being analyzed for which standard  preparations are
                not available.)  Transfer required amounts of  the appro-
                priate reference material into 100-ml volumetric  flasks
                using microliter pipets.  Dilute  to volume with Freon.
                The most appropriate pathlength for the quartz cells to
                be used in the spectrophotometric determination is
                determined by the expected sample concentration.  The
                following information is presented as a guide  to  selec-
                ting cells:
                      Pathlength, cm         Expected Range,  mg

                             1                        U - UO
                             5                      0.5-8
                            10                      0.1 - k
                Scan the standards  and samples from 3200  to 2700  cm 1
                using a recording infrared spectrophotometer.   Freon
                should be used in the reference beam or to zero the
                instrument.  The maximum absorbance at 2930 cm"1  should
                be used to construct a standard curve. The most  useful
                curve would be a plot of absorbance vs. mg oil as deter-
                mined by dilution of the standard reference solutions.

            b_.  Gravimetry. *  Transfer the water  extract  from  the col-
                lection vessel to a tared distillation flask.   Rinse the

                                 3-281

-------
                collection vessel with. Freon  and  add the washing to the
                distillation flask.   Distill  off  the solvent using a water
                bath at 70C.   After  the  solvent  has been  evaporated,
                place the flask on a  warm steam bath for 15 min.  During
                the final minute on the steam bath, draw air through the
                flask by means of an  applied  vacuum.   Cool the  flasks in
                a desiccator for 30 min and weigh.  The gain in weight
                of the tared flask is attributable to  oil  and grease if
                the Freon is free of  residue.
Calculations
            Select the appropriate method and  calculate the oil and
grease concentration based on the method of quantification that was used.
            a_.   Infrared spectrophotometry.  When  colorimetry  is used,
                prepare a standard curve by plotting measured  absorbance
                versus oil and grease concentration of the standards.
                Compare the absorbance of the  Freon extract to the
                standard curve to determine the  oil and grease concentra-
                tion.
                Calculate the oil and grease concentration  0  + G  of the
                original water sample as follows:
                                  o
                where
                     X = the concentration  of  oil  and  grease in the
                         Freon extract , mg/

                     V = the volume of Freon extract ,  H

                     S = the volume of sample  extracted,  H.  This is
                         determined by refilling sample collection
                         bottle to  the mark and measuring the required
                         volume in  liters.  This volume should be
                         corrected  for any  acid added  as  a preservative.
            b_.   Gravimetry .   When the amount of oil and grease is deter
                mined by weighing the material extracted, the sample
                concentration is determined as follows:

                               0 +  G mg/ = (A - B - C)
                where
                     A = weight of tared  flask  and  oil  and  grease
                         residue,  mg

                     B = weight of tared  flask, mg

                     C = calculated residue based on Freon  blank, mg

                     S = volume of water  initially  extracted, i.  This is
                         determined by refilling sample collection bottle

                                3-282

-------
to the mark and measuring the required volume
in liters.  This volume should "be corrected for
any acid added as a preservative.
         3-283

-------
                  Procedure for Sediment Samples (.SID)

            The procedure for determination of oil and grease in sedi-
ment samples is similar to that used to quantify oil and grease in
water samples.  The sample is extracted with Freon and the extractable
material is quantified.  As indicated in Figure 3-39 5 a moist sediment
sample must be used as a dried sample -will yield low results.  Because
of the operational definition of the oil and grease procedure and the
lack of precision associated with the test, it is recommended that con-
ditions of sampling, sample pretreatment, and analysis be standardized
to ensure comparability of the final data.
Method 1:  Freon Extraction
Apparatus
Extraction apparatus, Soxhlet
Vacuum pump or other source of vacuum
Extraction thimble, paper
Infrared spectrophotometer or balance
Reagents
Either cone, hydrochloric acid, HC1, or cone, sulfuric acid, HaSOit.
Magnesium sulfate monohydrate:  prepare MgSOu  HaO by drying a thin
      layer of MgSOi*  7^2 0 overnight in an oven at 103 C.
Freon (l,l,2-trichloro-l,2,2-trifluoroethane), boiling point klC.  The
      solvent should leave no measurable residue on evaporation.  Redi-
      still if necessary.
Grease-free cotton:  extract nonabsorbent cotton with Freon.
Known oil reference standard:  accurately weigh about 0.05 g of known
      oil directly into a 100-ml volumetric flask.  Add 80 ml Freon and
      dissolve the oil.  If, as in the case of a heavy fuel oil, all
      the oil does not go into solution, let stand overnight.  Filter
      through a Whatman No. hO filter paper into a second 100-ml volu-
      metric flask and dilute to volume with Freon.
Unknown oil reference standard (10 yl = 7-69 mg oil):  pipet 15 ml
      n-hexadecane, 15 ml isooctane, and 10 ml benzene into a 50-ml
      glass-stoppered bottle.  Assume the specific gravity of this
      mixture to be 0.769 and maintain the integrity of the mixture
      by keeping the bottle stoppered except when withdrawing aliquots.
Procedure
            Weigh a 20.0-g sample of moist sediment (SID) in a 150-ml

-------
"beaker.  (.The solids content of the sample should be known in advance or

determined on a separate sample aliquot.)  Acidify the sample with cone.

sulfuric or cone, hydrochloric acid to pH 2.

            Add 25 g MgSOii '  HzO to the acidified sediment sample.  Stir

to make a uniformly smooth paste that is spread on the beaker wall.

Allow to stand 15 to 30 min until solidified.  Remove the solids and

grind in a porcelain mortar.   The use of a desiccated, uniformly ground

sample improves the efficiency of the extraction process.

            Add the ground sample to a paper extraction thimble.  The
beaker and mortar should be wiped with a small piece of filter paper

that has been soaked in Freon.  Add the filter paper to the paper thimble.

Fill the thimble with glass wool or small glass beads.  Extract the pre-

pared sample with Freon in a Soxhlet apparatus at a rate of 20 cycles/hr

for k hr.  If the final extract is turbid, filter the sample through

grease-free cotton into a clean flask.  Rinse the initial sample con-

tainer and the cotton with Freon and add the washing to the filtered
sample.  Determine the oil and grease concentration of the extract by
either infrared spectrophotometry (a_) or gravimetry (b_).   The infrared

method would be preferred because it is generally more precise; partic-

ularly at low oil and grease  concentrations.
                                           1 9
            _a.  Infrared spectrophotometry.  '   Quantitatively transfer
                the sediment  extract to a convenient size volumetric
                flask and dilute to volume with Freon.
                Prepare calibration standards using either the known oil
                reference standard or the unknown oil reference standard.
                (.A known oil is defined as the only grease and/or oil
                component in the samples being analyzed.   An unknown
                oil is defined as the grease and/or oil component(s) in
                the sample being analyzed for which standard prepara-
                tions are not available.)  Transfer required amounts of
                the appropriate reference material into 100-ml volu-
                metric flasks using microliter pipettes.   Dilute to
                volume with Freon.
                The most appropriate pathlength for the quartz cells to
                be used in the spectrophotometric determination is
                determined by the expected sample concentration.  The
                following information is presented as a guide for
                selecting cell length:
                                 3-285

-------
                      Pathlength.,  cm          Expected Range, mg


                             5                      0.5-8
                            10                      0.1 - k

                Based on observed  ranges of oil and grease in sediments,
                it may be necessary to dilute the sample extracts  to  the
                working ranges indicated above.

                Scan the standards and samples from 3200 to 2700 cm"
                using a recording  infrared spectrophotometer.   Freon
                should be used in  the reference beam of a dual  beam
                instrument or to zero a single beam instrument.  The
                absorbance of the  2930-cm"1  peak should be used to con-
                struct a standard  curve.

            b_.  Gravimetry.l  The  gravimetric determination of  oil and
                grease does not require dilution of the samples.   However,
                the procedure is considered less precise than the  infra-
                red determination  because the method is subject to posi-
                tive sulfur interference and greater uncertainty at low
                oil and grease concentrations.

                To implement the method, quantitatively transfer the
                sediment extract to a tared distilling flask.   Rinse  the
                extract container  with Freon and add to the distilling
                flask.  Distill the Freon from the extraction flasks
                using a water bath at 70C.   After the solvent  has been
                evaporated, place  the flask on a warm steam bath for
                15 min and draw air through the flask by means  of  an
                applied vacuum for the final 1 min.   Cool in a  desiccator
                for 30 min and weigh.  The gain in weight is due to oil
                and grease if the  solvent is free of residue.

Calculations

            Select the appropriate method and calculate the oil and grease

concentration based on the method  of quantification that was used.
            eu  Infrared spectrophotometry.   When colorimetry is used,
                prepare a standard curve by plotting measured absorbance
                vs. oil and grease concentration of the standards.
                Compare the absorbance of the Freon extract to  the
                standard curve to  determine the oil and grease  concen-
                tration.

                Calculate the oil  and grease concentration  0 + G  of the
                original water sample as follows:

                        0 + GJ ing/kg (wet weight) =

                        0 + G mg/kg (dry weight) =
                                 3-286

-------
where
       X = concentration of oil and grease in the
           Freon extract, mg/
       V = volume of Freon extract , &

       g = wet weight of sediment extracted, g
     % S = percent solids in the sediment sample
           (expressed as a decimal fraction)

Gravimetry .  When the amount of oil and grease is
determined by weighing the material extracted, the
oil and grease concentration of the sediment sample
is calculated as follows:

     0 + G mg/kg (wet weight) = (A ~ B -
     0 + G mg/kg (dry weight) = (A "*> ()100)
where
       A = weight of tared flask and oil and grease
          . residue, mg
       B = weight of tared flask, mg
       C = calculated residue based on Freon flask, mg

       g = wet weight of sediment extracted, g
       S = percent solids in the sediment sample
           (expressed as a decimal fraction)
                 3-287

-------
                              References
1.  American Public Health Association.   Standard Methods for the Exami-
    nation of Water and Wastevater Including Bottom Sediments and
    Sludges. ll+th Edition.  APHA; New York, New York.   1193 p. (.1976).

2.  Environmental Protection Agency.   "Manual of Methods for Chemical
    Analysis of Water and Wastes."  Methods Development and Quality
    Assurance Research Laboratory, National Environmental Research
    Center; Cincinnati, Ohio.  298 p. (19710 .

3.  Disalvo, L., Guard, H., Hirsch, N.,  and Ng, J.  "Assessment and
    Significance of Sediment-Associated  Oil and Grease in Aquatic
    Environments."  Naval Biosciences Laboratory, Naval Supply Center;
    Oakland, California.  Technical Report D-77-26; U.  S. Army Engineer
    Waterways Experiment Station, CE; Vicksburg, Mississippi.  1^5 P
    (1977).
                                 3-288

-------
                       CHLORINATED HYDROCARBONS

            Chlorinated hydrocarbons are man-made compounds generally
used as insecticides or pesticides.  A second group of compounds, poly-
chlorinated biphenyls (PCB's), have also received extensive usage as
plasticizers.  Because of the hydrophobic nature of these compounds,
water concentrations are usually very low and sediment concentrations
are higher due to the combined processes of sorption and sedimentation.
            Chlorinated hydrocarbon residues must be concentrated prior
to analysis.  The methods available for sample concentration are solvent
                                 i*
extraction and carbon absorption.    However, because of a lack of con-
trol on the sorption-desorption processes, solvent extraction should be
considered the method of choice.  While 100 percent recovery would be
ideal, any solvent system that produces greater than 80 percent recovery
                         ry
is considered acceptable.   The solvent extraction procedure will concen-
trate chlorinated hydrocarbons for analysis but will also concentrate
interfering substances such as pesticide degradation products, pesticide
metabolites, lipid material, and, in the case of sediments, elemental
sulfur.3  Therefore, the extracts must be cleaned up prior to quantifi-
cation.

                      Sample Handling and Storage

            Samples for chlorinated hydrocarbon analysis should be
stored in glass bottles.2  To prevent sample contamination on the part
of the sample bottle caps or the cap liner, samples should be sealed with
either Teflon or acetone/hexane washed heavy-duty aluminum foil.
            Storage is less critical with chlorinated hydrocarbons than
with other organic compounds such as organophosphate and carbamates due
to the increased stability of chlorinated hydrocarbons.  Consequently,
sediment samples may be stored in a field moist, air-dried, or frozen
condition (Figure 3-^tO).   An increase in the chlorinated hydrocarbon
residue stability can be achieved by extracting the samples as soon as
possible to minimize the effects of microbial degradation.	
* References can be found on page 3-318.
                                 3-289

-------


AC 1 D 1 FY
WATER SAMPLE
4
4

FILTER
1
ACIDIFY
i 1
STORE
STORE
1 1
1 EXTRACT
EXTRACT
1 i
ANALYZE
(Wl)
ANALYZE
(W2)

f$ SAMPLE DESIGNATION
 PURPOSE












1
CORE SAMPLE
4

DREDGE SAMPLE
4 , -
r ^~
NO TREATMENT
(W,l STORE WET


f 4 T
_ 	 fc CLUTRIATC BIOASSAY

i i
ANALYZE ANALYZE
(SIA) (SID)

Wl W2 W3 SI A SIC


1
CORE SECTION
^
^
DRY
, 1
STORE
, 1
EXTRACT
i
ANALYZE
(S2)

SID 52

^^
FREEZE
1
STORE
i
EXTRACT
1
ANALYZE
(S3)


S3
Total Water Soluble Used in Mobile Bioavail- Total Total Total
Cone. Water Elutriate Cone. ability Sediment Sediment Sediment
Cone. Cone. Cone. Cone.
CONTAINER
SAMPLE TREATMENT
                            None
                                       Filter
                                                   None
                                                                None
                                                                                       None
                                                                                                   None      Ai r Dry      Freeze
PRESERVATIVE
                                                   None
                                                                  (Minimize Air Contact.   Keep Field Moist.)
                                                                                                             None
                                                                                                                         None
STORAGE TIME
                                                   
-------
            The use of plastic equipment and/or utensils during sample
collection, storage, and handling is to be avoided.


              Procedures for Water Samples (Wl, W2, SLA.)
Method 1:  Benzene Extraction"*

Apparatus

All glassware must be washed with heavy-duty soap and hot water and rinsed
      well.  Glassware should be rinsed with analytical grade acetone,
      pesticide grade ethyl acetate, and finally with sufficient quanti-
      ties of pesticide-grade hexane.  After rinsing, heat the glassware
      in an oven at 300 to ^00C overnight.  (.Heating along will not
      remove all the organic constituents.)  Rinse the glassware again
      with hexane prior to use.  For heavily contaminated glassware,
      such as those used to store concentrated pesticide standards,
      soaking in ethyl acetate may be required after the rinsing
      procedure.  The use of this glassware during sample analysis is
      to be avoided if at all possible

Gas chromatograph:  Varian 2800 or a Microtek 220, equipped with electron
      capture detector and recorder

Gas chromatograph columns:  the following columns have been used for the
      separation of chlorinated hydrocarbons:

      a.,  h percent SE-30 and 6 percent QF-1 on 60-80 (or 100-120) mesh
          size Chromosorb W, acid washed, DMCS treated

      b_.  6 percent QF-1 and k percent DC-11 on 100-120 mesh size on
          Chromosorb W, acid washed and DMCS treated
      c_.  11 percent OV-17/QF-1 (.commercially prepared by Applied Science
          Lab, Inc.) on 100-120 Gas-Chrom Q
      d_.  h percent OV-101/6 percent OV-210 on Chromosorb W, acid washed
          and HDMS treated, 80-100 mesh size

Disposable pipets

Graduated centrifuge tubes, 15 ml with glass stoppers

Volumetric flasks, 5, 10, and 100 ml

Round-bottomed flasks, 200, 300, and 500 ml, with 2^Ao ground-glass joint
Rotary evaporator

Chromatographic columns, 20 mm by ^00 cm, with Teflon stopcocks

Micro-syringes, Hamilton, 10 yl for injections, and other sizes such as
      25, 50, 100, and 250 yl for preparation of standard solutions

^00C oven
Oven for storing Florisil at 130C


                                3-291

-------
Muffle furnace capable of reaching 900C
Filter funnel, with 10 x k cm reservoir, porosity B sintered glass disc
      (.available from Ace Glass, Inc.)
Reagents
All solvents must be of pesticide grade and checked before use.   All
      chemicals must be of highest purity and, if applicable, should be
      preheated to eliminate artifacts or interferences.
Acetonitrite.
Acetone.
Hexane or petroleum ether.
Benzene.
Iso-Octane or toluene.
Chloroform.
Alumina, pretreated.
Anhydrous sodium sulfate, pretreated.
Florisil, 60-100 mesh, calcined at 650C (factory treated) and kept at
      130C until use.
Pesticide standards and standard solutions.
Procedure
            Sample pretreatment consists of three steps.   Chlorinated
hydrocarbons- and PCB's are isolated and concentrated by solvent extrac-
tion.  The extract is then subjected to successive cleanup on alumina
and Florisil columns.  Finally, the extract is analyzed using gas chroma-
tography.  At a minimum, the extract should be analyzed using two
columns of different polarity.
Extraction
            Add approximately 25 ml benzene to 1 5- of water sample in
the original sample bottle.  Stir the mixture for 30 min with a magnetic
stirrer so the vortex formed at the surface almost reaches the bottom
of the bottle.  (Wash the stirring bar in acetone and hexane prior to
use.)  Quantitatively transfer the mixture to a l- separatory funnel.
Rinse the sample bottle with two 30-ml benzene washes and .add to the
separatory funnel.
            Vigorously shake contents of separatory funnel and allow
organic layer to separate.  If an emulsion forms, add one-of the follo-
wing:  saturated sodium sulfate solution, methanol, isopropanol, or

                                 3-292

-------
2-octanol and gently agitate.
NOTE 1:  Added reagents should be checked to make sure that they do not
         contribute interferring peaks.
NOTE 2:  Alcohol addition should be limited to 5 to 10 drops to avoid
         a large, interferring solvent peak.
            Transfer the aqueous layer back to the empty sample bottle.
Dry the organic layer by rapid suction through 50 g sodium sulfate in
a filter funnel.  Store the organic extract in a 300-ml round-bottomed
flask.
            Add 25 ml benzene to the aqueous phase in the sample bottle.
Stir for 10 min and transfer to the separatory funnel.  Rinse sample
bottle with 20 ml benzene and add to separatory funnel.  Transfer the
aqueous layer to the sample bottle.  Dry the organic layer as before
and add to the first extract.
            Repeat the extraction process a third time using a 30-ml
portion of benzene.
            To the combined organic extracts, add 1 ml iso-octane and
concentrate on a rotary evaporator to approximately 3 ml.  During the
evaporation process, the water bath temperature should not exceed HoC.
When the extract has been concentrated to 10 to 12 ml, let the flask
rotate in air away from the water bath until the final volume of approxi-
mately 3 ml has been achieved.  This step is critical as severe loss
of some pesticides may occur if the water bath is too warm or the
extract is allowed to go to dryness.
Alumina cleanup
            Transfer the concentrated extract to a 15-ml graduated
centrifuge tube.  Rinse with, hexane and add to the centrifuge tube.
Evaporate the extract, under a gentle stream of nitrogen, to a volume
of approximately 1 ml.
            Prepare a microcolumn for sample cleanup by plugging cleaned
disposable pipets with a piece of precleaned glass wool.  Add 2 in.  of
deactivated alumina, prepared by mixing neutral alumina with 5 percent
of its weight of distilled water and tumbling for 2 hr before use.
Top the column with 1/2 in. of anhydrous sodium sulfate.
            Using a disposable pipet, quantitatively transfer the

                                3-293

-------
extract onto the column and wash the centrifuge tube with 1 ml 25 percent
benzene in hexane (.1:3 benzene:hexane).   As soon as the concentrated
extract sinks down to the sodium sulfate layer, transfer the benzene/
hexane washing to the column.  Wash the  tube with an additional 2 ml
of 25 percent benzene and transfer to the column.  After the solvent
sinks into the sodium sulfate layer, elute the column with 25 percent
benzene until 10 ml eluate is collected.   Add 0.5 ml iso-octane to the
eluate and evaporate to 0.5 ml in a 50C water bath under a gentle
stream of nitrogen gas.
            If PCB's are known to be absent and/or only a limited number
of chlorinated hydrocarbons are known to be present, this solution can
be diluted to volume and analyzed by gas chromatography.  If PCB's are
present and/or a complex mixture of chlorinated hydrocarbons is present
in the sample, the extract must be fractionated on a Florisil column
prior to quantification.
Florisil cleanup
            Fill a 20- by UOO-mm chromatographic column with a coarse
sintered disc near the bottom approximately three-fourths (3A) full
with hexane.  Add 2 g pretreated sodium  sulfate followed by 10 g of
Florisil added in portions.  Each portion should be 60-100 mesh, 650C
factory treated, stored at 130C, and cooled in a desiccator before
use.  Tap the column gently while adding the Florisil to the column  to
prevent channeling in the column.  Drain some hexane from the column
to settle the Florisil.  Add 3 g pretreated sodium sulfate to minimize
disturbance of the Florisil layer.
            Prewash the column with 50 ml benzene, followed by two
successive additions of 75 ml hexane. Allow the column to drain and
discard eluates.
            Dilute the concentrated sample extract to approximately  2 ml
with hexane.  Quantitatively transfer the sample to the column.  Allow
the extract to sink just to the surface  of the sodium sulfate layer.
Wash the round-bottomed flask with 3 ml  hexane and transfer the washing
solution to the column.  Let the extract run down as before.  Rinse  the
flask with two additional 3-ml hexane portions and add  each to the column.
                                3-29U

-------
Carefully add 100 ml hexane to the column without disturbing the Flori-
sil layer.
            Run the eluate into a 200-ml round-bottomed flask.  Place
sample on a roto-evaporator and concentrate to 10 to 12 ml in a ^0C
water bath.  Remove from the water bath and continue to rotate the
flask in the air until the volume is reduced to 3 ml.
NOTE 3:  Do not overheat sample or take to dryness.
            Quantitatively transfer the concentrate to a 15-ml graduated
centrifuge tube.  Wash the flask with 2 to 3 ml petroleum ether and add
to the centrifuge tube.  Repeat the rinsing procedure.  Add 0.5 ml iso-
octane (.or toluene) and concentrate sample to 0.5 ml under a gentle
stream of nitrogen.  This fraction is ready for GC analysis.  (See NOTE
U.)
            Elute the same column with 100 ml 6 percent ethyl ether in
petroleum ether (.or 6 percent ethyl ether in hexane).  Catch the eluate
in a clean, 200-ml round-bottomed flask.   Concentrate the eluate as
above.  This fraction is ready for GC analysis.   (See NOTE U.)
            Elute the column a third time with 100 ml 15 percent ethyl
ether in petroleum ether (.or 15 percent ethyl ether in hexane).  Catch
eluate in a clean, round-bottomed flask and concentrate as before.
This fraction is ready for GC analysis.  (See NOTE k.)
            Repeat column extraction with 100 ml 50 percent ethyl ether
in petroleum ether (.or chloroform) and proceed as above.  The final
extract is ready for GC analysis.  (.See NOTE k.)
NOTE 4:  The four fractions can be analyzed for the following chlorinated
         hydrocarbons:
         Fraction 1 (.hexane) - ^BHC, heptachlor, aldrin, p,p*-DDE,  and
                               PCB's (Aroclor 1248, 125^, and 1260).
         Fraction 2 (6 percent ethyl ether in hexane) - p,p1-DDD,
                               pjp^DDT,  Ojp^DDT, lindane, "-chlordane,
                               transchlordane, methoxychlor, and hepta-
                               chlor epoxide.
         Fraction 3 (-15 percent ethyl ether in hexane) - endrin,
                               ^-endosulfur, and dieldrin.
         Fraction 4 (chloroform) - 3-endosulfan.
                                3-295

-------
            Identification of chlorinated hydrocarbon pesticides and
PCB's should be based on retention time on at least two different
columns of different polarity (Table 3-20).   Confirmation is based on
the preparation and identification of chemical derivatives,  thin-layer
chromatography, and/or mass spectroscopy.
Calculations
            A standard calibration curve should be prepared  daily.
Pesticide concentrations are determined by comparing the sample reponse
to the standard curve (.provided the recorder response is less than
TO percent of full scale and the peak height [or area] is close to that
of the standard) as follows:
             g chlorinated hydrocarbon/^ = /^\/^\/\ * 10~3
where
     A = peak height (or area) produced by sample
     B = picograms standard injected into GC
     C = final volume of sample concentrate, ml
     D = peak height (.or area) produced by B
     E = volume of water initially extracted, H
     F = volume of sample extract injected to produce A,  yl
Remarks
            Factory-calcined Florisil (at 650 C)  varies in activity from
batch to batch.  It is necessary to standardize a new batch when it is
received; the activity should be checked periodically to ensure it  does
not change upon storage.  A large batch of Florisil should be subdivided
quickly into smaller portions (a portion is taken out and then subdivided)
in a dehumidified room and each portion stored in a tightly capped  brown
bottle in a desiccator.  Enough supply of Florisil for a week or so is
transferred in a glass- stoppered bottle and kept  at 130C until used.
NOTE:  Do not unnecessarily expose Florisil to the atmosphere.
            Standardization of Florisil:   use a mixed nanogram solution
containing 10 ng/Ul each of lindane, heptachlor,  aldrin,  heptachlor
epoxide, p^-DDE, 20 ng/yl of p^-DDD and ko ng/yl of p.p^DDT.
Prepare a Florisil column containing 10 g Florisil as described earlier.
                                3-296

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                                                 Table 3-20

                   Retention Times of Various Organochlorine Pesticides Relative to Aldrin
1>J



Pesticide
ccBHC
PCNB
Lindane
Dichloran
Heptachlor
Aldrin
Heptachlor
epoxide
Endosulfan I
p.p^DDE
Dieldrin
Captan
Endrin
o^-DDT
p,pa-DDD
Endosulfan II
p.p^DDT
Mirex
Methoxychlor
Aldrin
(min absolute)
Relative Retention Time Under
Liquid Phase:
1.5* OV-17 +
+1.95* QF-1T
0.51*
0.68
0.69
0.77
0.82
1.00

1.5U
1.95
2.23
2.UO
2.59
2.93
3.16
3.U8
3.59
U.18
6.1
7.6

3.5
Given Conditions*

Liquid Phase:
5% OV-210tt
0.6U
0.85
0.81
1.29
0.87
1.00

1.93
2.U8
2.0
3.00
U.09
3.56
2.70
3.75
^.59
U.07
3.78
6.5

2.6
     * All columns glass, 180 cm by U mm I.D.,  solid support  Gas-Chrom Q (100/120 mesh).
    ,T Column temperature, 200C; argon/methane carrier  flow, 60 ml/min.
       Column temperature, l80C; argon/methane carrier  flow, 70 ml/min.

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Dilute 50 yl of the mixed standard to approximately 1 ml in a tube and
transfer the solution to the column.  Follow Florisil Elution Procedure
to obtain three fractions.  Concentrate each fraction to 10 ml and
examine by GLC.
            The elution rate should be adjusted to 5 to 6 ml/min.   The
first fraction (hexane or petroleum ether)  should contain:   heptachlor,
aldrin, and DDE.   The second fraction should contain:  lindane,  hepta-
chlor epoxide, DDD, and DDT; and the last fraction (.15 percent ether
in hexane or petroleum ether) should contain dieldrin.  If the separa-
tion is not clear cut (i.e-. overlapping of the fractions),  Florisil
can be increased or decreased first by 2 g, then narrowed down to  1 g.
Alternatively, volume of elution solvent can also be adjusted to obtain
complete fractionation; however, this approach is limited only to  minor
overlapping of different fractions.
            If separation of DDE anS PCB's is desirable, the charcoal
column developed by Berg et al.   can be used.

Method 2:  Methylene Chloride/Hexane Extraction2
Apparatus
Gas chromatograph fitted with electron capture, flame photometric, and
      electrolytic conductivity  detectors
Gas chromatograph columns, 22 by 300 mm, with Teflon stopcocks,  without
      frits.  Use one of the following column packing mixtures:
      a_.  1.5 percent OV-17/1.95 percent OV-210 - Liquid phases  premixed
          and coated on silanized support,  80-100 mesh size
          Instrument operating conditions with this column are:  operating
          temperature, 200C; detector temperature, 205C;  carrier flow,
          50-70 ml/min
      b_.  h percent SE-30/6 percent OV-210 - Liquid phases  premixed and
          coated on silanized support, 80-100 mesh size
          Instrument operating conditions with this column are:  opera-
          ting temperature, 200C; detector temperature, 205C;  carrier
          flow, 70-90 ml/min
      c_.  5 percent OV-210 - coated on silanized support, 100-120  mesh
          Instrument operating conditions with this column are:  opera-
          ting temperature, 200 C; detector temperature, 205 C;  carrier
          flow, ^5-60 ml/min
                                3-298

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Water bath capable of maintaining 95 to 10CP C
Separatory funnels, 2 &, with Teflon stopcocks
Filter tubes, 150 by 2U mm, Corning 9^80 or the equivalent
Kuderna-Danish concentrator fitted with graduated evaporative con-
      centrator tube.  These are available from the Kontes Glass
      Company, each component bearing the following stock numbers:
      a_.  Flask, 500 ml, Stock # K-570001
      b_.  Snyder column, 3-ball, Stock # K-503000
      c_.  Steel springs, 1/2 in., Stock # K-662750
      d_.  Concentrator tubes, 10 ml, Size 1025, Stock # K-570050
Modified micro-Snyder columns, 19/22, Kontes Stock # K-569251
Glass beads, 3 mm, plain, Fisher Stock # 11-312 or equivalent
Modified micro-Snyder column, 19/22 T-joint, Kontes Stock # 569251
Pipet, vol., h ml
Reagents
Hexane, pesticide quality, distilled in glass.
Isooctane, pesticide quality.
Diethyl ether, AR grade,  peroxide free.  The ether must contain 2 percent
      (.v/v) absolute ethanol.  Most of the AR grade ethyl ether contains
      2 percent ethanol, added as a stabilizer; it is, therefore,
      unnecessary to add ethanol unless it is found necessary to remove
      peroxides.
NOTE:  To determine the absence of peroxides, test in accordance with the
       method outlined elsewhere.
Petroleum ether, pesticide quality, redistilled in glass; b.p. 30 to
      6oc.
Methylene chloride, pesticide quality.
Methylene chloride/hexane, 15 percent v/v.
Eluting mixture, 6 percent (.6 + 9^):  60 ml of diethyl ether is diluted
      to 1000 ml with petroleum ether and approximately 15 g of anhy-
      drous Na2SOit is added to ensure freedom from moisture.
Eluting mixture, 15 percent (.15 + 85):   150 ml of diethyl ether is
      diluted to 1000 ml with petroleum ether and approximately 15 g
      of anhydrous NaaSOi^ is added.
NOTE:  None of the eluting mixtures should be held longer than 2U hr
       after mixing.
Anhydrous sodium sulfate, reagent grade, granular, Mallinkrodt Stock
      # 802h or equivalent.
                                3-299

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NOTE:  The purity of this material should "be tested as outlined elsewhere
       (Section 5, A, (..l), P3)2  except that 15 percent methylene chlo-
       ride/hexane should he substituted for petroleum ether.
Florisil, 60/100 mesh, PR grade.
Procedure
            It is assumed that final thin layer chromatography and elec-
trolytic conductivity confirmation may be applied to supplement the
information obtained by electron capture detection.  For this  reason,
a larger sample is used than would be necessary for electron capture
detection alone.  Dilution of an aliquot of the final extract  for
analysis using gas chromatography with electron capture detection
requires less time than the extraction of a second sample for  identity
confirmation.
            Transfer 2  of sample (or a lesser volume, if indicated)
to a h-H separatory funnel and add 120 ml of 15 percent methylene
chloride/hexane (MC/hexane).
NOTES:  If, on the basis of prior analysis of a given waterway, the
        residue levels may be expected to run high, a sample of 500 ml
        or 1  may be indicated.  In this event, the size of the
        separatory funnel should be 2  and the extraction solvent
        volumes given as 120 ml should be reduced to 100 ml.
        A 500-ml graduated cylinder is a suitable measuring device for
        the initial sample.  Any measuring discrepancy up to 5.0 ml
        would result in an error no greater than 1.0 percent.
            Stopper funnel and shake vigorously for 2 min.  Allow layers to
separate and draw off aqueous layer into a second 2- separatory funnel.
            Add another 120 ml of 15 percent MC/hexane to the  aqueous
phase in separatory funnel #2, stopper, and shake vigorously for another
2 min.
            Prepare a 2-in. column of anhydrous, granular NaaSOi, in a
150- by 2U-mm filter tube with a small wad of preextracted glass wool  at
the bottom.  Position this over a 500-ml K-D flask to which is attached
a 10-ml concentrator tube with one 3-mm glass bead in the bottom.
            Filter the MC/hexane extract in the separatory funnel #1
through, the NaaSOij column into the flask.
            Draw off the aqueous layer in separatory funnel #2 into
empty separatory funnel #1.

                                 3-300

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            Add 120 ml of straight hexane to the aqueous solution in
separatory funnel #1, stopper, and shake again for 2 min.   Draw off  and
discard the aqueous layer.
            Filter the solvent extracts in both separatory funnels through
the NaaSOi* into the flask, rinsing down filter tube with three 10-ml
portions of hexane.
            Attach a 3-ball Snyder column to the K-D flask, place
assembly in a boiling water bath, and concentrate extract  to approxi-
mately 5 ml.
            Remove K-D assembly from bath, cool, and rinse T-joint be-
tween tube and flask with a small volume of hexane; also rinse down
walls of tube.  Rinse should be delivered with 2 ml Mohr pipet and
should not exceed 3 ml.
            Place tube under a slow nitrogen stream at ambient tempera-
ture and reduce extract volume to approximately 0.5 ml. Using a dis-
posable pipet, carefully add hexane to adjust volume to exactly 1.0  ml
in the tube tip.  Then, with a U-ml vol. pipet, add k ml of hexane.
Do not rely on the accuracy of the tube graduation at the  5-ml mark.
            Stopper concentrator tube and mix vigorously on a vortex
mixer for 1 min.  The sample is now ready for GLC analysis.
            It should be possible to make some tentative compound
identifications upon computation of relative retention times (RRT) of
peaks in the preliminary chromatograms via electron capture.  Full
reliance should not be placed on the chromatographic data  obtained from
one column.  An alternate column of completely different compound
elution characteristics should be used to (.a) confirm a number of com-
pounds tentatively identified on the first column and (b)  isolate and
tentatively identify any compound pairs which may have eluted as single
peaks on the first column.
            If the initial chromatogram indicates the presence of a
sufficient amount of interfering materials, it may prove necessary to
conduct a Florisil cleanup on the extract.  Based on general experience,
this is rarely necessary on most surface water samples.  If it should
prove necessary, process the cleanup as discussed on the next page.
                                3-301

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Extract cleanup procedure
            Prepare a chromatographic column containing U in.  (.after
settling) of activated Florisil topped with 0.5 in. of anhydrous
granular NaaSOt,.  A small vad of glas-s wool, preextracted with petro-
leum ether, is placed at the "bottom of the column to retain the
Florisil.
NOTES:  If the oven is of sufficient size, the columns may be  prepacked
        and stored in the oven, withdrawing columns a few minutes "before
        use.
        The amount of Florisil needed for proper elution should be
        determined for each lot of Florisil.
            Place a 500-ml Erlenmeyer flask under the column and prewet
the packing with petroleum ether (1*0 to 50 ml, or a sufficient volume
to completely cover the Na2SOit layer).
NOTE:  From this point and through the elution process, the solvent
       level should never be allowed to go below the top of the Na2SOit
       layer.  If air is introduced, channeling may occur, resulting in
       an inefficient column separation.
            Using a 5-ml Mohr or a long disposable pipet, immediately
transfer the extract (.approximately 5 ml) from the evaporator  tube onto
the column and permit it to percolate through.
            Rinse tube with two successive 5-ml portions of petroleum
ether, carefully transferring each portion to the column with  the pipet.
NOTE:  Use of the Mohr or disposable pipet to deliver the extract
       directly onto the column precludes the need to rinse down the
       sides of the column.
            Prepare two Kuderna-Danish evaporative assemblies  complete
with 10-ml graduated evaporative concentrator.tubes.  Place one glass
bead in each concentrator tube.
            Replace the 500-ml Erlenmeyer flask under each column with
a 500-ml Kuderna-Danish assembly and commence elution with 200 ml of
6 percent diethyl ether in petroleum ether (Fraction l).  The  elution
rate should be 5 ml per min.  When the last of the eluting solvent
reaches the top of the Na2SOit layer, place a second 500-ml Kuderna-
Danish assembly under the column and continue elution with 200 ml of
15 percent diethyl ether in petroleum ether (Fraction 2).
            To the second fraction only, add 1.0 ml of hexane  containing
                                3-302

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200 nanograms of aldrin, place both Kuderna-Danish evaporator  assemblies
in a water bath, and concentrate extract until approximately 5 ml remain
in the tube.
            Remove assemblies from bath and cool to ambient  temperature.
            Disconnect collection tube from Kuderna-Danish flask and
carefully rinse joint with a little hexane.
            Attach modified micro-Snyder column to collection  tubes,
place tubes back in water bath, and concentrate extracts  to  1  ml.   If
preferred, this may be done at room temperature under  a stream of nitro-
gen.
            Remove from bath and cool to ambient temperature.  Disconnect
tubes and rinse joints with a little hexane.
NOTE:  The extent of dilution or concentration of the  extract  at this
       point is dependent on the pesticide concentration  in  the substrate
       being analyzed and the sensitivity and linear range of  the
       electron capture detector being used in the analysis.
            Should it prove necessary to conduct further  cleanup on the
15 percent fraction, transfer 10 g MgO-Celite mixture  to  a chromato-
graphic column using vacuum to pack.   Prewash with approximately hd ml
petroleum ether, discard prewash, and place a Kuderna-Danish receiver
under column.  Transfer concentrated Florisil eluate to column using
small portions of petroleum ether.  Force sample and washings  into  the
MgO-Celite mixture by slight air pressure and elute column with 100 ml
petroleum ether.  Concentrate to a suitable volume and proceed with gas
liquid chromatography.
NOTE:  Standard recoveries should be made through column  to  ensure
       quantitative recoveries.
            Inject 5 yl of each fraction into the gas  chromatograph for
the purpose of determining the final dilution.   If all peaks are on
scale and quantifiable, it will not be necessary to proceed  with any
further adjustment in concentration.
            If off-scale peaks are obtained in either  fraction, it  will
be necessary to dilute volumetrically with hexane to obtain  a  concentra-
tion that will permit quantification of those peaks from  a 5-  to 10-yl
injection.
            If the electron capture data indicate the  probable presence
                                3-303

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of one or more chlorinated pesticide compounds,  the chromatographer
would be veil advised to conduct confirmation via electrolytic  conduc-
tivity detection in the reductive mode even though positive  identifi-
cations were made on two columns via electron capture.   This extra step
provides needed validation, particularly when compounds  are  tentatively
identified which appear to be out of place in light of known supple-
mental data concerning the waterway sampled.
            It is improbable that parent compounds in the  organo-
phosphorus class will be detected in an average  water sample.   Compound
degradation is rather rapid in the aqueous medium.   However, if the
waterway receives heavy runoff from nearby agricultural  land undergoing
current spray programs, the presence of these residuals  is possible.
Calculations
            A standard calibration curve should  be prepared  daily.
Pesticide concentrations are determined by comparing the sample response
to the standard curve (provided the recorder response is less than
TO percent of full scale and the peak height [or area! is  close to that
of the standard) as follows:
            ug chlorinated hydrocarbon/^ = 'W\\  x 10~3
where
     A = peak height (or area)  produced by sample
     B = amount of standard injected into GC,  picograms
     C = final volume of sample concentrate, ml
     D = peak height (.or area)  produced by standard B
     E = volume of water initially extracted,  
     F = volume of sample extract injected to  produce  A, yl
Remarks
            This method will not detect the acid form  of herbicides  such
as 2,U-D or 2,H,5-T, but should be suitable for certain of the  esters of
these compounds which are used  commercially.   However, as these compounds
are only about one tenth (.1/10) as responsive  to electron capture as a
number of the common chlorinated pesticides, it appears somewhat remote
that they would be detected in  an average water sample by this  procedure.
            In a laboratory study conducted on river water in the Water

                                3-30U

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                                                                        2
Quality Laboratory of the Environmental Protection Agency in Cincinnati,
the degradation pattern shown in Table 3-21 was reported on a 20-gal
sample of water held in the laboratory under sunlight and fluorescent
light.  Thepe data are presented for supplemental information.
            The two fractions from the Florisil column should never be
combined for examination by gas liquid chromatography.  By so doing,
a valuable identification tool is voided.
            Meticulous cleaning of glassware is absolutely essential for
success with this procedure.  All reagents and solvents must be pretested
to ensure that they are free of contamination by electron capturing
materials at the highest extract concentration levels.  Reagent blanks
should be run with each set of samples.
            The method, as described, is known to be capable of pro-
ducing recoveries of most of the chlorinated pesticides of from 85 to
100 percent.  Each laboratory should conduct its own recovery studies
to make certain of its capability to achieve this recovery range.  A
clue may be obtained from the recovery of the aldrin spike.   The recovery
of this compound should not be less than 70 percent.
            For the removal of peroxides from the ethyl ether,  place an
appropriate volume in a separatory funnel and wash it twice with portions
of water equal to about one half the volume of ether.  The washed ether
is shaken with 50 to 100 ml of saturated NaCl solution and all  of the
aqueous layer is discarded.  The ether is then transferred to a flask
containing a large excess of anhydrous sodium sulfate and shaken vigorously
on a mechanical shaker for 15 min.  This treatment should not be attemp-
ted on ether-containing ethanol, as the amount of ethanol that  would
remain is indeterminate.
            If the presence of malathion is suspected, it is necessary
to pass 200 ml of 50 percent diethyl ether in petroleum ether through
the Florisil column into a third K-D evaporator assembly, concentrating
the eluate as described for the 6 percent and 15 percent eluates.
                                3-305

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                           Table 3-21
       Persistence  of  Chlorinated Hydrocarbon Pesticides in
River Water


Compound
Organochlorine
Compounds
BHC
Heptachlor
Aldrin
Heptachlor
epoxide
Telodrin
Endosulfan
Dieldrin
DDE
DDT
DDD
Chlordane (.tech.)
Endrin
Organopho sphorus
Compounds
Parathion
Methyl parathion
Malathion
Ethion
Trithion
Fenthion
Dimethoate
Merphos
Merphos recov.
as Def
Azodrin
Car "hamate
Compounds
Sevin
Zectran
Matacil
Mesurol
Baygon
Monuron
Fenuron
in Terms


0-time


100
100
100

100
100
100
100
100
100
100
100
100


100
80
100
100
90
100
100
0

100
100


90
100
100
90
100
80
80
of Percentage Recovery2

Original
1 wk


100
25
100

100
25
30
100
100
100
100
90
100


50
25
25
90
25
50
100
0

50
100


5
15
60
0
50
1*0
60



Compound Found* , %
2 wk


100
0
80

100
10
5
100
100
100
100
85
100


30
10
10
75
10
10
85
0

30
100


0
0
10
0
30
30
20
U wk


100
0
hO

100
0
0
100
100
100
100
85
100


< 5
0
0
50
0
0
75
0

10
100


0
0
0
0
10
20
0
8 wk


100
0
Ho

100
0
0
100
100
100
100
85
100


0
0
0
50
0
0
50
0

< 5
100


0
0
0
0
5
0
0
 Pesticide concentrations were 10 ug/&.  Recoveries were rounded
off to the nearest 5  percent.

                              3-306

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            Procedures for Sediment Samples (SID, S2, S3)2

Method 1:  Acetone/Hexane Extraction
            The examination of sediment from the bottom of a stream or
lake provides information concerning the degree of contamination resul-
ting from pesticides, particularly the organochlorine compounds which
are not readily biodegradable.  This information, combined with residue
data obtained by analysis of the water and tissues from resident marine
life, contributes to the development of an overall profile of the pesti-
cidal contamination of a given body of water.
            The sediment sample is partially dried and extracted by
column elution with a mixture of 1:1 acetone/hexane.   The extract is
washed with water to remove the acetone and then the pesticides are
extracted from the water with 15 percent CH2Cla in hexane.   The extract
is dehydrated, concentrated to a suitable volume, subjected to Florisil
partitioning, desulfurized, if necessary, and analyzed by gas chroma-
tography.
Apparatus
Gas chromatograph:  Varian 2800, Microtek 220, or equivalent, equipped
      with an electron capture detector and recorder
Gas chromatograph columns, 22 by 300 mm, with Teflon stopcocks
Gas chromatograph column packing  (See Methods 1 and 2 for the analysis
      of water.)
Pans, approximately lU by 10 by 2-1/2 in.
Oven, drying
Muffle furnace
Desiccator
Crucibles, porcelain, squat form, Size 2
Omni or Sorvall mixer with chamber of approximately ^00 ml
Separatorv funnels, 500 and 250 ml, with Teflon stopcocks
Filter tube, 180 by 25 mm
Kuderna-Danish concentrator fitted with graduated evaporative concentrator
      tube.  Available from the Kontes Glass Company, each component
      bearing the following stock numbers:
      a..  Flask, 250 ml, Stock # K-570001
      b.  Snyder column, 3-ball, Stock # K-503000
                                 3-307

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      .  Steel springs, 1/2 in., Stock # K-662750
      d..  Concentrator tubes, 10 ml, Size 1025, Stock # K-570050
Pyrex glass wool, preextracted with methylene chloride in a Soxhlet
      extractor
Hot water bath, temperature controllable at 8CPC
Reagents
Sodium sulfate, anhydrous, Baker, prerinsed or Soxhlet extracted with
      methylene chloride.
n-Hexane, pesticide quality.
Acetone, pesticide quality.
Methylene chloride, pesticide quality.
Ac etone-hexane, 1:1
Diethyl ether, pesticide quality, free of peroxides
Distilled water, suitable for pesticide residue analysis
Sodium sulfate solution, saturated
Methylene chloride-hexane, 15 percent v/v
Procedure
            Decant and discard the water layer over the sediment.  Mix
the sediment to obtain as homogeneous a sample as possible and transfer
to a pan to partially air dry for about 3 days at ambient temperatures.
NOTE:  Drying time varies considerably depending on soil type and
       drying conditions.  Sandy soil will sufficiently dry in 1 day
       whereas muck requires at least 3 days.  The silt and muck sediment
       is sufficiently dry when the surface starts to split, but there
       should be no dry spots.   Moisture content will be 50 to 80 percent
       at this point.
            Weigh 50 g of the partially dried sample into a UoO-ml Omni
mixer chamber.  Add 50 g of anhydrous sodium s.ulfate and mix well with
a large spatula.  Allow to stand with occasional stirring for approxi-
mately 1 hr.
NOTE:  If the final calculations will be made on a dry basis, it is
       necessary at this point to initiate the test for percent total
       solids in the sample being extracted for pesticide evaluation.
       Immediately after weighing the 50-g sample for extraction,
       weigh approximately 5 g of the partially dried sediment into a
       tared crucible.  Determine the percent solids by drying overnight
       at 103C.  Allow to cool in a desiccator for half an hour before
       weighing.  Determine the percent volatile solids by placing the
       oven-dried sample into a muffle furnace and igniting at 550C
       for 60 min.  Allow to cool in a desiccator before weighing.
                                 3-308

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            Attach the UoO-ml chamber to an Omni or Sorvall mixer and
blend for about 20 sec.  The sample should be fairly free flowing at this
point.
            Carefully transfer the sample to a chromatographic column.
Rinse the mixer chamber with small portions of hexane, adding the rinsings
to the column.
            Elute the column with 250 ml of 1:1 acetone-hexane at a flow
rate of 3 to 5 ml/min into a UoO-ml beaker.
            Concentrate the sample extract to about 100 ml under a nitro-
gen stream and at a temperature no higher than 55C.  Transfer to a 500-
ml separatory funnel containing 300 ml of distilled water and 25 ml of
saturated sodium sulfate solution.  Shake the separatory funnel for
2 min.
            Drain the water layer into a clean beaker and the hexane
layer into a clean, 250-ml separatory funnel.
            Transfer the water layer back into the 500-ml separatory
funnel and reextract with 20 ml of 15 percent methylene chloride in
hexane, again shaking the separatory funnel for 2 min.  Allow the layers
to separate.  Discard the water layer and combine the solvent extracts
in the 250-ml separatory funnel.
            Wash the combined solvent extract by shaking with 100 ml of
distilled water for 30 sec.  Discard the wash water and rewash the extract
with an additional 10 ml of distilled water, again discarding the wash
water.
            Attach a 10-ml evaporator concentrator tube to a 250-ml
Kuderna-Danish flask and place under a filter comprised of a small wad
of glass wool and approximately 0.5 in. of anhydrous NaaSOij in a filter
tube.
            Pass the solvent extract through the drying filter into the
K-D flask, rinsing with three portions of approximately 5 ml each of
hexane.
            Attach Snyder column to top joint of a K-D flask, immerse
tube in 80C water bath, and concentrate extract to 5 ml.
            Remove tube, rinsing joint with small volume of hexane.  The
sample is now ready for Florisil partitioning.
                                 3-309

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            Prepare a Florisil chromatographic column containing
h in.  (after settling) of activated Florisil topped with 0.5 in. of
anhydrous, granular NaaSOij.  A small wad of glass wool, preextracted
with hexane, is placed at the "bottom of the column to retain the
Florisil.
NOTES:  If the oven is of sufficient size, the columns may "be prepacked
        and stored in the oven, withdrawing columns a few minutes
        before use.
        The amount of Florisil needed for proper elation should be
        determined for each lot of Florisil.
            Place a 500-ml Erlenmeyer flask under the column and
prewet the packing with hexane (^0 to 50 ml, or a sufficient volume
to completely cover the Na2SOit layer).
NOTE:  From this point and through the elution process, the solvent
       level should never be allowed to go below the top of the
       NaaSOij layer.  If air is introduced, channelling may occur,
       making for an inefficient column.
            Assemble two more K-D apparatus but with 500-ml flasks
and position the flask of one assembly under the Florisil column.
However, at this point, use 25-ml graduated evaporator concentrator
tubes instead of the 10-ml size for previous concentrations.
            Using a 5-ml Mohr or a long disposable pipet, immediately
transfer the extract from the evaporator tube onto the column and
permit it to percolate through.  Rinse tube with two successive
5-ml portions of hexane, carefully transferring each portion to the
column with the pipet.
NOTE:  Use of the Mohr or disposable pipet to deliver the extract
       directly onto the column precludes the need to rinse down
       sides of the column.
            Commence elution with 200 ml of 6 percent diethyl ether
in petroleum ether (Fraction l).  The elution rate should be approxi-
mately 5 ml per min.  When the last of the eluting solvent reaches
a point approximately 1/8 in. from the top of the NaaSOi* layer,
place the second 500-ml Kuderna-Danish assembly under the column
and continue elution with 200 ml of 15 percent diethyl ether in
petroleum ether (Fraction 2).  Place both Kuderna-Danish evaporator
assemblies in a water bath and concentrate extract to approximately
20 ml.
                                3-310

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NOTE:  If there is reason to suspect the presence of malathion in the
       sample, have a third 500-ml K-D assembly ready.  At the end of
       the 15 percent fraction elution, add 200 ml or 50 percent
       diethyl ether in petroleum ether (Fraction 3), evaporating the
       eluate in the same manner.
            Remove K-D assemblies from the bath, cool, and rinse the
T-joint between the tube and flask with a little petroleum ether.
Finally, dilute both extracts to exactly 25 ml and proceed with the
GLC determinative step.
            Inject 5 yl of each fraction extract into the gas chroma-
tograph (.electron capture mode) primarily to determine whether the
extracts will require further adjustment by dilution or concentration.
            When appropriate dilution adjustments have been made in
the extracts and the column oven is set at the required temperature,
the relative retention values of the peaks on the chromatograms
should be calculated.  When these values are compared with the values
in Table 3-20 for the appropriate column, the operator should be able
to make tentative compound identifications.  Microcoulometry and/or
TLC may be required for positive confirmation of some of the suspect
chlorinated compounds, whereas flame photometric detectors (FPD)  may
be utilized for the organophosphate suspects.
            An analytical problem that must be considered when
sediment samples are analyzed for chlorinated hydrocarbon pesticides
is sulfur interference.  Elemental sulfur is encountered in most
sediment samples, marine algae, and some industrial wastes.  The
solubility of sulfur in various solvents is very similar to the
organochlorine and organophosphate pesticides; therefore, the sulfur
interference follows along with the pesticides through the normal
extraction and cleanup techniques.  The sulfur will be quite evident
in gas chromatograms obtained from electron capture detectors,
flame photometric detectors operated in the sulfur or phosphorus
mode, and Coulson electrolytic conductivity detectors.  If the gas
chromatograph is operated at the normal conditions for pesticide
analysis, the sulfur interference can completely mask the region
from the solvent peak through aldrin.
            This technique eliminates sulfur by the formation of

                                3-311

-------
copper sulfide on the surface of the copper.   There are two  critical
steps that must be followed to remove all the sulfur:   (a) the copper
must be highly reactive--therefore, all oxides must be removed so that
the copper has a shiny, bright appearance; and (b)  the sample extract
must be vigorously agitated with the reactive copper for at  least
1 min.
            It will probably be necessary to  treat  both the  6 and
15 percent Florisil eluates with copper if sulfur crystallizes out
upon concentration of the 6 percent eluate.
            Certain pesticides will also be degraded by this tech-
nique, such as the organophosphates, chlorobenzilate,  and heptachlor.
However, these pesticides are not likely to be found in routine
sediment samples because they are readily degraded  in the aquatic
environment.
            If the presence of sulfur is indicated  by an exploratory
injection from the final extract concentrate  (presumably 5 ml) into
the gas chromatograph, proceed with removal as follows:
            a_.  Under a nitrogen stream at ambient  temperature,
                concentrate the extract in the concentrator
                tube to exactly 1.0 ml.
            b_.  If the sulfur concentration is such that crystal-
                lization occurs, carefully transfer, by syringe,
                500 yl of the supernatant extract (or  a lesser
                volume if the sulfur deposit  is too heavy) into a
                glass-stoppered, 12-ml graduated conical centrifuge
                tube.  Add 500 yl of iso-octone. Add approximately
                2 g of bright copper powder,  stopper,  and mix
                vigorously 1 min on a vortex  genie  mixer.
                NOTE:  The copper powder, as  received from the
                       supplier, must be treated for removal of
                       surface oxides with 6  ITHNOa.  After  about
                       30 sec of exposure, decant acid and rinse
                       several times with distilled water and
                       finally with acetone.   Dry under a nitrogen
                       stream.
            c_.  Carefully transfer 500 yl of  the supernatant-treated
                extract into a 10-ml graduated concentrator  tube.
                An exploratory injection into the gas  chromatograph
                at this point will provide information as to whether
                further quantitative dilution of the extract is
                required.
                               3-312

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Calculations
            The chlorinated hydrocarbon pesticide concentration of
sediment can be calculated as:
        Chlorinated hydrocarbons yg/kg (vet weight) = )EWF\/r.\

     Chlorinated hydrocarbons yg/kg (dry weight) = (E)(F)(G)(g s)
where
       A = nanograms standard injected into GC
       B = peak height (or area) produced by sample injection
       C = final volume of sample extract, ml
       E = peak height (or area) produced by standard injection A
       F = wet weight of sediment sample initially extracted, g
       G = volume of extract injected to produce B, ml
     %, S = sediment percent solids as a decimal fraction

Method 2:  Acetonitrile Extraction1*
            The sediment is extracted with acetonitrile and the
chlorinated hydrocarbons are partitioned into petroleum ether.
The ether extract is cleaned up on a Florisil column and separated
into four fractions for subsequent GLC analysis.  This method has
been used to quantify the following chlorinated hydrocarbons and
PCB's (.with the value in parenthesis being the lowest level of
detection in ppm):
      Lindane                                          (O.OOl)
      Heptachlor                                       (O.OOl)
      Heptachlor Epoxide                               (O.OOl)
      Aldrin                                           (O.OOl)
      Dieldrin                                         (O.OOl)
      p,p'-DDD                                         (0.001)
      p,p'-DDT                                         (0.001)
      p,p'-DDE                                         (0.001)
      o,p'-DDT                                         (O.OOl)
      Endrin                                           (O.Ol)
      p,p'-methoxychlor                                (0.0 5)
      Ot-endosulfan                                     CO.Ol)
      3-endosulfan                                     (.0.01)
      cis-chlordane                                    (.0.005)
      trans-chlordane                                  (0.005)
      Aroclor 12k&                                     (0.100)
      Aroclor 1251*                                     (0.100
      Aroclor 1260                                     (0.100)
                               3-313

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Apparatus
Gas chromatograph, Varian 2800, Microtek MT220 or equivalent,  equipped
      with a flame photometric detector
GLC column packing materials  (See Methods 1 and 2 for the analysis of
      water.)
GLC columns  (See Methods 1 and 2 for the analysis of water.)
Disposable pipets
Magnetic stirrer and 5/8-in. Teflon-coated stirring bars
Graduated centrifuge tubes, 15 ml with glass stoppers
Kuderna-Danish (K-D) evaporator and associated glassware
Volumetric flasks
Hamilton micro-syringes, 10 yl for GLC injection and 100 yl for pre-
      paration of standard solutions
Graduated pipets, 2 and 10 ml
Heating plate
Vortex genie
Reagents
All solvents must be of pesticide grade.  All chemicals must be of
      highest purity and, if applicable, should be pretreated  to
      eliminate artifacts or interferences.
Acetonitrile.
Hexane or petroleum ether.
Benzene.
Diethyl ether containing 2 percent ethanol as preservative.
Florisil, 60-100 mesh, calcined at 650C (factory treated) and kept
      at 130C until used.  (See discussions in the procedure  for
      the analysis of chlorinated hydrocarbon pesticides and PCB's
      in water, Methods 1 and 2.)
Anhydrous sodium sulfate, pretreated.
Neutral alumina, Woelm, activity Grade I deactivated with 5 percent
      water.  (See Method 1 for analysis of water.)
Pesticide standard and standard solutions.
Procedure
            Quantification of chlorinated hydrocarbon pesticides is
a three-step procedure consisting of extraction, extract cleanup,
and identification.
            Extraction.  Transfer 10 g dry weight equivalent of
sediment into the glass jar of a Waring blender with a Bakelite top.

-------
(.Do not use a rubber or plastic top.)  Add 120 ml of acetonitrile and
blend at medium-high speed for 15 min.  Allow solid particles to settle
somewhat.  Pour the acetonitrile extract, which may contain some sus-
pended particles, into an Allihn filter tube containing prewashed
celite covering the sintered glass.
NOTE:  If the residue in the Allihn filter tube becomes excessive, it
       should be scraped out with a spoon-type spatula and combined
       with that in the blender before the second blending and extrac-
       tion.
            To the residue in the blender, add another 120 ml of
acetonitrile and hQ ml of distilled water and blend for 10 min.
Filter as before.
            Pour 60 ml of acetonitrile into the blender and blend the
homogenate for 10 min.  Transfer all the residue, if necessary,  with
2 by 20 ml acetonitrile, into the Allihn tube and filter.   Apply
strong suction so that the residue in the tube contains little
solvent.
            Transfer, with petroleum ether rinsing, the combined
acetonitrile extracts into a l- funnel and dilute with distilled
water to adjust the aqueous content to 20 percent.  Extract the
resulting mixture with 150 ml and then twice with 75 ml petroleum
ether.
            Wash the combined petroleum ether extracts with
approximately 200 ml distilled water.  Discard water washing and
pass the organic extract under suction or with air pressure, through
an anhydrous sodium sulfate (10 to 15 g) column using a 500-ml
round-bottomed flask as a receiver.
            In a rotary evaporator, evaporate the contents in the
500-ml flask to 2 or 3 ml.  (Do not let contents get dry and do  not
use a water bath temperature over UoC for evaporation; otherwise,
there will be possible loss of pesticides and PCB's.  See  evapora-
tion precautions discussed in the procedure for water analysis.)
            Cleanup.  Transfer the concentrated petroleum  ether
extract with a clean disposable pipet onto a 30-g (.for the exact
amount to be determined, see procedure for water analysis) Florisil
column with 0.5 in. of anhydrous sodium sulfate on the top of the
                                 3-315

-------
Florisil.  Use a 300-ml round-bottomed flask as  a receiver.
            Allow the extract to sink down just  to the sodium sulfate
layer.  Rinse the round-bottomed flask with 2 or 3 ml of petroleum ether
and transfer the rinsing with the same disposable pipet onto the column.
Let the rinsing solvent again sink down just to  the sodium sulfate layer.
Rinse the round-bottomed flask again with 2 or 3 ml of petroleum ether
and transfer the rinsing onto the column.
            Again rinse the round-bottomed flask, this time  with 20 to
30 ml petroleum ether.  Carefully pour the petroleum ether onto the
column so that the sodium sulfate layer is not disturbed.  Elute the
column with a total of 200 ml (including the above rinsings) of
petroleum ether.
            Concentrate eluate with a rotary evaporator to 1 or 2 ml
and transfer, with benzene rinsings, to a 10-ml  volumetric flask.
Make up to 10 ml with benzene for GLC examination (Fraction l).
            Change the receiver and elute column with 200 ml of
6 percent diethyl ether containing 2 percent ethanol.  Concentrate
eluate to 1 to 2 ml on a rotary evaporator.  Transfer to a 10-ml
volumetric flask.  Rinse round-bottomed flask with benzene and  add to
volumetric flask.  Dilute to volume with benzene.  This fraction,
Fraction 2, is now ready for GLC analysis.
            With a third 300-ml round-bottomed flask as receiver,
elute the column with 200 ml of 15 percent ether in petroleum ether.
Concentrate to 10 to 20 ml with a rotary evaporator.  Add 50 to
60 ml of benzene and concentrate to 1 to 2 ml.  Make up to 10 ml with
benzene in a volumetric flask (Fraction 3).
            Elute the column with 200 ml chloroform or 200 ml
50 percent diethyl ether in petroleum ether.  Collect the eluate
in a round-bottomed flask and concentrate to 2 to 3 ml on a  rotary
evaporator.  Add 50 to 60 ml benzene and reduce  the volume to 2 to
3 ml.  Add a second 50- to 60-ml portion of benzene and evaporate
to 2 to 3 ml.  Transfer the concentrate to a 10-ml volumetric flask;
Rinse the round-bottomed flask with benzene and  add the rinsing to
the volumetric flask.  This fraction is now ready for GLC analysis
(Fraction U).
                                 3-316

-------
            The petroleum ether fraction (Fraction l)  contains PCB's,
heptachlor, aldrin, p,p'-DDE, and ^-BHC.
            The 6 percent diethyl ether-petroleum ether fraction
(Fraction 2) contains lindane, heptachlor epoxide, p,p'-DDT,  p,p'-DDD,
methoxychlor, o,p'-DDT5 cis-chlordanes, and trans-chlordanes.
            The 15 percent diethyl ether-petroleum fraction (Fraction  3)
contains dieldrin, K-endosulfan, and endrin.
            The last fraction (.k) contains 3-endosulfan.
            Extracts may have to be cleaned up for sulfur interference.
Follow procedures given in Sediment Method 1.
            Identification.  Examine the above four eluates by GLC.
Further concentration and dilution may be necessary to produce on-
scale GLC peaks.  Procedures for confirmation  of identity are the
same as for water extracts.
Calculations
            The concentration of chlorinated hydrocarbon pesticides
in the sediment samples can be calculated as follows:
        Chlorinated hydrocarbons yg/kg (wet weight) =  ;  (/  \ >(

     Chlorinated hydrocarbons yg/kg (dry weight) = '(E)(F)(HW g)
where
       A = weight of picograms of standard injected
       B = peak height (or area) of sample
       C = volume of sample extract, ml
       E = peak height (or area) of standard
       F = volume of sample extract required to produce B,  yl
       H = wet weight of sediment initially extracted, g
     % S = percent solids in sediment (expressed as a  decimal fraction)
                                 3-317

-------
                              References
1.  Brodtman,  N.  V.,  Jr.   "Quantitation of Chlorinated Pesticides  
    A Comparison of Methods."  JAWWA:558-560 (1975).
2.  Thompson,  J.  F. (.Ed.)   "Analysis of Pesticide Residues  in Human
    and Environmental Samples."  Environmental Protection Agency
    Pesticides and Toxic  Substances Effects Laboratory; Research
    Triangle Park, North Carolina (197*0 .
3-  Jensen, S., Renberg,  L.,  and Reutergardh,  L.   "Residue  Analysis
    of Sediment and Sewage Sludge for  Organochlorines  in the
    Presence of Elemental  Sulfur."  Anal.  Chem. 1*9:316-318  (1977).
k.  Environment Canada.   "Analytical Methods Manual."   Inland Waters
    Directorate,  Water Quality Branch; Ottawa, Ontario, Canada (197*0
5.  Berg, 0. W.,  Diosady,  P.  L., and Rees, G.  A.  V.  Bull.  Environ.
    Contam. Toxic, 7, 338.
                                3-318

-------
                     ORGANOPHOSPHOROUS PESTICIDES

            Organophosphates have received usage as pesticides  because
                                                      i*
of their ability to inhibit the enzyme cholinesterase.     They  are
generally more acutely toxic than the compounds they vere designed to
replace, organochlorine hydrocarbons, but they also degrade more
rapidly.  The methods to quantitate organophosphorous compounds in
environmental samples are based on organic extraction followed  by
gas chromatographic analysis2 or an enzyme inhibition method.1
Fenitrothion is determined by hexane extraction; dimethoate and
phosphamidon are determined by chloroform extraction;2  while 1^
other organophosphorous can be determined by benzene extraction.2
Because a phosphorous-specific detector is used, only minimal sample
cleanup is generally required.  The second method does not provide
a quantitation of specific organophosphorous pesticides.   Rather,
the method provides an estimate of all compounds that can deactivate
an enzyme.  Thus, the results will include carbamates as  well as
organophosphate pesticides that are present in the sample.

                      Sample Handling and Storage

            Samples should be collected and stored in clean glass
containers.  Because of the reactivity of organophosphates, samples
should be extracted in the field whenever possible.   At other times,
samples should be maintained at UC and extracted as soon as possible.
There is no known acceptable storage period for samples but extended
residue stability is obtained by extraction.  Also,  several organo-
phosphates (dimethoate, phosphamidon, fenitrothion)  may degrade in
the presence of sunlight.2  Samples should only be exposed to sub-
dued lighting to minimize this potential effect on sample integrity.
Only wet sediment samples should be collected as suggested in
Figure 3-^1.
*  References can be found on page 3-336.
                                3-319

-------
Figure 3-Ul.    Handling and  storage of samples for organophosphate analysis
                                                         CORE SAMPLE

*
*
WATER SAMPLE DREDGE SAMPLE CORE SECTION
i * i
4 4- * ^
STORE FILTER N TENT STORE WET
1 1 	
EXTRACT STORE
1
EXTRACT

I I \
f ELUTRIATE BIOASSAY FXTRACT

11 1
ANALYZE ANALYZE ANALYZE ANALYZE
(Wl) (W2) (S1A) (SID)
U)
ro SAMPLE DESIGNATION
O
PURPOSE
CONTAINER
SAMPLE TREATMENT
PRESERVATIVE
STORAGE TIME
DIGESTION SOLUTION
SAMPLE VOLUME OR WEIGHT
Wl W2 W3 S1A SIC SID
Total Water Soluble Used in Mobile Bioavail- Total
Cone. Water Elutriate Cone. ability Sediment
Cone. Cone.
G G G G G G
None Filter None None None None
I*C ltC None 
-------
               Procedure for Water Samples (Wl, W2, S1A)2

Method 1:  Hexane, Chloroform, Benzene Extraction
            The procedure consists of three extractions on tvo sample
aliquots.  One aliquot is extracted with hexane to recover fenitro-
thion.  The residual aqueous phase or solid residue is extracted with
chloroform to recover phosphamidon and dimethoate.  A second sample
aliquot is extracted with benzene to recover lU other orthophosphates.
Apparatus
Gas chromatograph:  Varian 2800, MicroTek MT 220, or equivalent,
      equipped with a flame photometric detector, with independent
      power supply, electrometer, and 1.0-mV dual pen recorder
Gas chromatograph columns:  three columns are specified.   However,
      column a_ is most useful "because the packing, consisting of
      OV-1T + QF-1, separates all lU organophosphorous pesticides
      under isothermal analysis at 200C.  The other columns do
      not separate all residues but may be used for partial con-
      firmation:
      a_.  11 percent (w/w) OV-17 + QF-1 on Chromosorb Q,  80-100 mesh
          (available from Applied Science Laboratory, Inc.)
      b_.  3.6 percent OV-101 and 5-5 percent OV-210 on Chromosorb W,
          80-100 mesh, acid washed and DMCS treated
      c_.  3 percent OV-225 on Chromosorb W HP, 80-100 mesh
Gas chromatograph operating conditions:  all analyses are conducted
      under isothermal column conditions.  Other operating tempera-
      tures are:  injection port, 210C; column temperature, 200 C; and
      detector temperature, l85C.  Gas flows should be optimized
      for maximum sensitivity with parathion.  Suggested values are
      nitrogen, 80 ml/min; hydrogen, 150 ml/min; oxygen,  20 ml/min;
      and air, 10 ml/min
Disposable pipets
Magnetic stirrer and 5/8-in. Teflon-coated stirring bar
Kuderna-Danish (K-D) evaporator and associated glassware
Graduated centrifuge tubes, 15 ml with glass stoppers
Volumetric flasks, 10 ml
Hamilton micro-syringes, 10 yl for sample injection and 100 yl for
      preparation of standard solutions
Graduated pipets, 2 and 10 ml
Beaker, 250 ml for water bath
Heating plate
                                 3-321

-------
Vortex genie mixer

Rotary evaporator and associated equipment

Round-bottomed flasks, 500 and 300 ml

Separatory funnel, 2 

Suction funnel to fit 500-ml round-bottomed flask

Nitrogen gas, prepurified or better

Reagents

All solvents must be of pesticide residue grade.

Hexane.

Chloroform.

Benzene.

Acetone.

Toluene.

Anhydrous sodium sulfate.

Analytical standards, obtainable from manufacturers.   Working solu-
      tions (for injection) should be prepared veekly.  The con-
      centration of this standard should be chosen for one-half
      full-scale deflection.  (See preparation of standard solutions.)
Preparation of standard solutions

Solvents:  benzene or ethyl acetate are acceptable solvents for the
      preparation of standard solutions.   Benzene is  appropriate
      for the working solutions that are injected into the GLC.

Stock solution:  in a 100-ml low actinic volumetric flask dissolve
      100 mg of pure, analytical grade pesticide  standard in a few
      milliliters of ethyl acetate and make up to volume with
      benzene.  Shake flask well and keep in a refrigerator when
      not in use.

Working stock solution A:  from the individual stock  solutions at
      room temperature, pipet the following volumes of each pesti-
      cide listed below and transfer to a clean,  10-ml volumetric
      flask.  Dilute to the 10-ml mark with benzene.   Shake flask
      well and keep in a refrigerator when not in use.

                           1.0 ml diazinon
                           1.0 ml parathion
                           1.0 ml ethion
                           1.0 ml ronnel
                           1.0 ml malathion
                           2.0 ml methyl trithion
                           0.5 ml disyston

Working stock solution B:  from the individual stock  solutions at
      room temperature, pipet the following volumes of each pesticide
      listed on the.next page and transfer to a clean, 10-ml volumetric

                                3-322

-------
      flask.  Dilute to the 10-ml mark with benzene.   Shake  well  and
      keep in a refrigerator when not in use.

                           1.0 ml methyl parathion
                           0.5 ml thimet
                           2.0 ml trithion
                           2.0 ml ruelene

Nanagram solution (l.O ng/Ul):   from the working stock solutions  A and
      B, and the following stock solutions:  imidan,  guthion,  and
      ethyl guthion, withdraw 100 yl of each and transfer  to a clean,
      10-ml volumetric flask and dilute to the 10-ml  mark  with benzene.
      Shake well.
      The resulting concentration of each pesticide standard in the
      nanogram solution is given in Table 3-22.

                                 Table 3-22
              Composition of Organophosphorous Nanogram Standard




Individual
Stock Solution
Azinphosethyl
(Ethyl Guthion )
Azinphosmethyl
(GuthionR)
Carbophenothion
(TrithionR)
Cruf ornate
(Ruelene^)
Diazinon
Disulfoton
(DisystonR)
Ethion
Imidan
Malathion
Methyl Parathion^
Methyl TrithionR
Parathion
Phorate
(ThimetR)
Ronnel
(TroleneR)
Amount
Withdrawn
f
-------
      Dilute to 10-ml mark with benzene.   Shake well.
Procedure
            The procedure consists of three extractions on two  aliquots
of the water sample.  One aliquot is successively extracted with hexane
and chloroform to recover fenitrothion and phosphamidon,  respectively.
A second aliquot is then extracted with benzene to recover the  remain-
ing organophosphorous compounds.
            Extraction of fenitrothion.  Stir the sample on a magnetic
stirrer so that the vortex formed at the  surface almost reaches the
bottom of the bottle.  While stirring, add 25 to 50 ml hexane and 100 g
sodium sulphate and tightly cover the bottle.
            After stirring for ^5 min, transfer the contents to a 2-Jl
separatory funnel.  Shake 2 min.  Transfer the aqueous layer back to
the sample bottle.  Dry the hexane layer  under rapid suction through
a short column of anhydrous sodium sulphate, into a 300-ml round-
bottomed flask and wash the column with 15 to 20 ml hexane.
            Repeat the hexane extraction  two more times using 25 to
50 ml of solvent each time.  Collect and  combine the extracts as
before.
            Concentrate the hexane on a rotary evaporator to less
than 5 ml (>0 to 50C water bath) .
            Quantitatively tranfer the hexane to a 15-ml centrifuge
tube using a few milliliters hexane.  Make up to 10 ml with hexane.
Shake well or place on vortex genie for 30 sec.  Analyze by GLC.
If response is low, transfer the extract  to a graduated centrifuge
tube.  Concentrate the sample to 0.5 ml in a 50C water bath, also
using a gentle stream of nitrogen.  Reanalyze the concentrate.
            Extraction of phosphamidon and dimethoate.  To the
aqueous solution from the first extraction add 25 to 50 ml chloroform
and stir on a magnetic stirrer for 20 min.  Transfer contents of
sample bottle to a 2- separatory funnel.  Shake for 2 min.
            Dry the chloroform layer by passing through a short
column of anhydrous sodium sulphate into  a 500-ml round-bottomed
flask and wash the column with chloroform.
            Rinse sample bottle with 25 to 50 ml of chloroform  and
                                3-32U

-------
transfer to separatory funnel containing aqueous solution.   Shake  2 min
and dry chloroform layer as before.   Repeat the extraction  a third time.
            Concentrate the chloroform on a rotary evaporator (^0  to
50C water bath) to less than 5 ml.
            Quantitatively transfer  the chloroform to a 15-ml centri-
fuge tube using a few millilters of  benzene.  Make up to 10 ml with
benzene.
            Add 0.5 nil toluene to centrifuge tube and concentrate  to
0.5 ml on kO to 50C water bath and using a gentle stream  of nitrogen.
Make up to 10 ml with benzene.
            Examine sample by GLC.  If response is low, transfer to a
15-ml graduated centrifuge tube.  Place sample in a 50C water bath
and concentrate to 0.5 ml under a gentle stream of nitrogen.   Reexamine
by GLC.
            Qualitative identification is based on relative retention
time of parathion on two different colums using the phosphorous
flame photometric detector.  The sulfur mode may also be used to
confirm an organophosphorous residue containing a sulfur atom.
Phosphamidon contains no sulfur atom.
            Extraction of other organophosphates.   A fresh  sample  is
extracted with benzene to quantify the lU organophosphates  listed  in
Table 3-23.
            Stir the sample on a magnetic stirrer so that the vortex
formed at the surface almost reaches the bottom of the bottle. While
stirring, add 10 ml benzene and 100  g sodium sulfate and tightly cap
the bottle.  After stirring 1*5 min,  let the layers separate and bring
the organic phase into the neck of the bottle by addition of distilled
water.
            If an emulsion occurs, it can be broken up by mechanical
stirring with a disposable pipet or  by addition of some sodium
sulfate.  Water-soluble, polar organic solvents are not recommended
because organophosphorous compounds  will partition into these
solvents and depressed recoveries are observed.
                               3-325

-------
                              Table 3-23
               Detection Limit for 1^ Organophosphorous
                   Pesticides in 1-& Water Samples*


                 Pesticide
      Azinphosethyl (Ethyl GuthionR)
      Azinphosmethyl (Guthion^)
      Carbophenothion (Trithion^)
      Crufornate (.Ruelene^)
      Diazinon
      Disulfoton (DisystonR)
      Ethion
      Imidan
      Malathion
      Methyl Parathion
      Methyl Trithion^
      Parathion
      Phorate (.ThimetR)
      Ronnel (TroleneR)
      *  One liter water extracted with 10 ml benzene and 5 ml
        extract concentrated to 0.5 ml.  Eight-microliter injec-
        tion on GLC at 2.56 x io~  amp full scale.   Detection
        limit detsTrrnned at about twice the noise.	
            Concentrate the extract by one of the following methods:
            a_.  Remove as much of the benzene extract as  possible
                and place in a 15-ml centrifuge  tube.  Record
                exact  volume.   Add 0.5 ml toluene,  place  in 60C
                water  bath, and blow a gentle stream of nitrogen
                over the extract, concentrating  it  to 0.5 ml.
                Make up volume (0.5 to 1.0 ml) with benzene and
                place  on a vortex mixer for 10 sec  to ensure  homo-
                geneity.   Examine the sample by  GLC.
            b_.  Remove as much of the benzene extract as  possible
                and place in a 10-ml K-D tube.  Record exact  volume.
                Add 0.5 ml toluene and an ebullator and place a
                3-ball Snyder condenser on K-D tube.   Place in K-D
                block  and wrap aluminum foil around condenser.
                Concentrate at a block temperature  of 130C to
                0.5 ml (about 30 min).  The K-D  block must be
                preheated.

                Make up to volume (0.5 to 1.0 ml) with benzene and
                place  on a vortex mixer for 10 sec  to ensure  homo-
                geneity.   Examine the sample by  GLC.
            Qualitative identification is based  on  relative retention

time to parathion on two different columns using the phosphorous

                               3-326

-------
flame photometric detector.  The sulfur mode may also be used to confirm
those organophosphorous compounds with a sulfur atom in the structure.
Calculations
            Peak height or area is employed to estimate concentrations  of
the organophosphorous residues in the water sample.   The flame photometric
detector is linear between 0.1 and 10 ng/injection for all these organo-
phosphorous residues except ruelene.  The calculations are as follows:
                 Organophosphorous yg/A = j^) (G)(H)
where
     A = nanograms of standard injected into GC
     B = peak height (or area) of sample
     C = volume of sample extract, ml
     D = final volume of extract after concentration, ml
     E = peak height (or area) of standard
     F = volume of extract injected into GC to produce response B, yl
     G = volume of water sample initially extracted,  
     H = volume of extract removed for concentration, ml
Confirmation of identity
            Two chemical derivatization techniques,  using chromous
chloride  and pentafluorobenzyl bromide,1* respectively, for the confir-
mation of organophosphorous pesticide residues have  been developed.
            Both methods have merit.  The CrCl2 method is comparatively
faster; but it is applicable to only three organophosphorous compounds.
The pentafluorobenzyl ether method is applicable to  eight compounds
which were investigated.  It is, therefore, more comprehensive;  but  it
takes comparatively longer than the CrCla method.
Additional information
            Cleanup is often unnecessary when the FPD is used,  as there
are low background interferences.  High concentration of sulphur in  the
extract, however, may lead to cross-channel interference and affect  the
response of the P-mode.
            The FPD is highly specific for P- and/or S-containing
compounds.
            Phosphamidon exists as a mixture of isomers, the a-trans

                                  3-327

-------
and B-eis in an approximate proportion of 27:73.   Since the  3-cis  isomer
is the largest, it was chosen for use in the peak height quantification
method.5
            On most columns examined (see Tables  3-2k  and 3-25)5 feni-
trothion and 3-phosphamidon had similar retention times and  could  not be
resolved.  Because of the solubilities of these organophosphorous  com-
pounds in various solvent systems, partitioning will successfully  sepa-
rate these two compounds.
            Phosphamidon and dimethoate are water soluble and  do not
partition into the hexane phase of a hexane-water system.  Fenitrothion
is quantitatively recovered in the hexane.
            Benzene will extract fenitrothion quantitatively but will
also recover some of the phosphamidon and dimethoate.
            A polar solvent will successfully extract  the water-soluble
phosphamidon and dimethoate from the aqueous solution.   It was found that
chloroform was the best solvent for quantitative  results.  Methylene
chloride was more soluble in water (2 g/100) and  depressed recoveries
were observed.
            The temperature of the water bath for concentrating extracts
is critical for consistent quantitative recoveries.  A temperature of
kOC is the best for recovery, but 50C is more practical for  time.
            If the CHCla extract is injected in the flame photometric
detector, there is no difference in peak height from that of a benzene
extract containing phosphamidon and dimethoate.  However,  when the CHCla
is vented off, anomalous and extraneous peaks may be observed  on the
electron capture detectors on the same instrument.  Therefore, the
chloroform is replaced by benzene for gas chromatographic analysis.
            The use of sodium sulfate was beneficial in both the
extraction of fenitrothion with hexane and phosphamidon and  dimethoate
with chloroform.
            Ruelene response is nonlinear above 8.0 ng/injection.
            The CrCla reduction of nitro group on fenitrothion is
applicable as a confirmatory test for fenitrothion.  This reaction
had no effect on either phosphamidon or dimethoate, even when  used
with ethylene diamine.  Fenitrothion may also be  confirmed as  the  PFB
ether.
                                  3-328

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                                             Table 3-2U
                                                                                    3
                Retention Time and Peak Height Data for Organophosphorous Pesticides




Pesticide
Phorate (ThimetR)
Diazinon
P
Disulfoton (Di-Syston )
p
Ronnel (Trolene )
Methyl Parathion
u>
^ Malathion
ro
^ Parathion
p
Cruf ornate (Ruelene )
Methyl TrithionR
Ethion
p
Carbophenothion (Trithion )
T ., R
Imidan
Azinphosmethyl (Guthion )
Azinphosethyl (Ethyl Guthion )


Concentration
of Standard
ng/u&
0.5
1.0
0.5
1.0
1.0

1.0

1.0
2.0
2.0
1.0
2.0
10.0
10.0
10.0
**
Average
Peak
Height
cm
15.11*
ll*.3l*
10.86
13.53
9.69

8.0k

9.75
5.17
6.10
6.00
6.kk
11. 2U
6.32
7.80
Coefficient
of Variation
of Peak
Height
. %
2.73
2.39
2.52
2.67
1*.31

3.98

1*.33
15. 2U
5.13
2.1*2
2.6l
7.17
7.1*6
3.32
t
Relative
Retention
Time t
min
0.28
0.3U
0.1*1
0.59
0.79

0.88

1.00
1.21
2.07
2.32
2.50
6.00
8.05
9.81
Coefficient
of Variation
of t^
R
%
0.57
0.56
0.63
0.27
O.l6

0.08

0.00
0.87
0.2U
0.22
0.25
0.2k
0.32
0.33
 *  Column packed with OV-17/QF-1.  Instrument operated iosthermally at 200C, attenuation 2.56 x 10  , amp
  full scale.
*  Average of ten determinations with an 8-yl injection.
   tp = 1.00 for parathion (6.9 min).

-------
                              Table 3-25


            Retention Time for Organophosphorous Compounds


                       on OV-101/OV-210 at 200C
                                                                 t  **
          Compound	                R.T.*                R	
Dimethoate                                  0.85                 0.5**





Aminofenitrothion                           1.05                 0.6T





a-Phosphamidon                              1.20                 0.79





Fenitrothion                                1.35                 0.86





Parathion                                   1.55                 1.00





Fenitrothion-oxygen analogue                1-55                 1.01





6-Phosphamidon                              1.65                 1.06
 *   Retention time in arbitrary units.

**  Relative retention time to parathion (tR parathion - 1.00).   These

   values were determined using an integrator as a timer.
                                  3-330

-------
            The hexane extraction recovers all lk organophosphorous com-
pounds with an 87 to 99 percent  recovery at 10 ppb, with the exception of
ruelene (30 percent).   The  retention time of ruelene does not interfere
with those of phosphamidon  or  dimethoate.
                                  3-331

-------
                  Procedure for Sediment Samples (SID)

Method 1:  Hexane Extraction2'6
Apparatus
Gas chromatograph:  Varian 2800, Microtek 220, or equivalent, equipped
      with a flame photometric detector and a recorder
Gas chromatograph columns:  three columns are specified.   Column a_ has
      the widest general applicability:
      a.  11 percent (.W/W) OV-17 + QF-1 on Chromosorb Q,  80-100 mesh
          (available from Applied Science Laboratory, Inc.)
      b_.  3.6 percent OV-101 and 5-5 percent OV-210 on Chromosorb W,
          80-100 mesh, acid washed and DMCS treated
      c_.  3 percent OV-225 on Chromosorb W HP, 80-100 mesh
Gas chromatograph. operating conditions:  all analyses are conducted under
      isothermal column conditions.  Other operating temperatures are
      injection port, 210 C; column temperature, 200C; detector tempera-
      ture, l85C.  Suggested gas flows are nitrogen, 80 ml/min; hydro-
      gen, 150 ml/min; oxygen, 20 ml/min; and air, 10 ml/min.
Disposable pipets
Magnetic stirrer and 5/8-in. Teflon-coated stirring bar
Kuderna-Danish (K-D) evaporator and associated glassware
Graduated centrifuge tubes>:  15 ml with glass stoppers
Volumetric flasks, 10 ml
Hamilton micro-syringes:  10 yl for sample injections and 100 yl for
      standard preparation
Graduated pipets, 2 and 10 ml
Beaker, 250 ml for water bath
Heating plate
Vortex genie mixer
Rotary evaporator and associated equipment
Round-bottomed flasks, 300 and 500 ml
Separatory funnel, 2 
Suction funnel to fit 500-ml round-bottomed flask
Nitrogen gas, prepurified or better
Reagents
All reagents must be of pesticide grade quality.
Hexane.
Chloroform.
                                  3-332

-------
Benzene.
Acetone.
Toluene.
Anhydrous sodium sulfate.
Analytical standards:  obtainable from manufacturers.   Working solutions
      should be prepared weekly and working standards  should be prepared
      daily.  The concentration of these standards should be chosen for
      one-half full-scale deflection.   Prepare standards according to
      directions provided in Method 1:  Procedures for Water Samples.
      Standards should be stored in dark bottles and refrigerated.
Procedures
            The sediment extraction procedure is similar to that used
for water samples.   Samples are sequentially extracted with hexane and
chloroform.  However, because of the reactivity of organophosphorous
compounds, only wet sediment samples (SID)  should be considered for
analysi s.
            Extraction of fenitrothion.  Weigh 50 g wet sediment sample
(SID) and transfer to a 250-ml Erlenmeyer flask.  Add  100 ml hexane
and 10 ml deionized distilled water.  Seal  the flask and shake for
15 min.
            Separate and retain the hexane  layer.  Repeat the extraction
two more times.  Combine the extracts and save the sediment slurry for
chloroform extractions.
NOTE:  Water is added to promote the desorption of pesticide residues
       from the sediment.  The extraction efficiency can also be
       improved with an ultrasonic homogenizer.
            Dry the hexane extract by passing through  an anhydrous
sodium sulfate column into the flask of a rotary evaporator.   Wash the
column with 15 to 20 ml hexane and add to the extract.   Concentrate
the extract to approximately 5 ml on a rotary evaporator in a ^0  to
50C water bath.
            Quantitatively transfer the concentrate to a 15-ml centri-
fuge tube using hexane to wash the round-bottomed flask.   Dilute to
volume with hexane.
            Mix the sample and inject a 10-ul aliquot  into the gas
chromatograph.  Adjust the concentrate volume as necessary by evapora-
tion or dilution to bring the response on scale.  Reanalyze the
                                  3-333

-------
concentrate.
            Extraction of phosphamidon and dimethoate.   Extract  the
sediment slurry aqueous extract with 100 ml chloroform.   Shake for 15 min
and separate the solvent layer.  Repeat the chloroform  extraction a
second and a third time and combine the extracts.
            Dry the extract by passing through an  anhydrous  sodium
sulfate column.  Wash the column with 15 to 20 ml  chloroform.  Combine
the column washing and the sample extract in a round-bottomed flask.
Reduce the volume to less than 5 ml using a rotary evaporator and a  kO
to 50C water bath.
            Transfer the extract to a 15-ml centrifuge  tube  and  add
0.5 ml toluene.  Concentrate to approximately 0.5  ml in a hcPC water
bath under a stream of nitrogen gas.  Dilute to 10 ml with benzene and
analyze by gas chromatography.
            Extraction of other organophosphates.   Extract a separate
50-g aliquot of the SID sample with 100 ml of benzene.   Shake for
15 min and separate the organic layer.  Repeat the extraction two more
times.
            Combine the extracts and dry in an anhydrous sodium  sulfate
column.  Transfer the extract to a round-bottomed  flask.   Reduce the
volume to approximately 5 ml using a rotary evaporator  and a 50C
water bath.
            Quantitatively transfer the concentrate to  a graduated
centrifuge tube and add 0.5 ml toluene.  Reduce the volume to 0.5 ml
in  a 50C water bath.  Dilute to volume with benzene and analyze by
gas chromatography.
Calculations
            Peak height or area is employed to estimate concentrations
of the organophosphorous residues in the sediment  sample.  The calcu-
lations are as follows:
       Organophosphates yg/kg (wet weight) = 	(E)(F)(.H)(g)	

       Organophosphates yg/kg (dry weight) = (E)(F)(H)(g)($  S)
                                  3-33^

-------
where
       A = nanograms of standard injected into GC
       B = peak height (or area) of sample response
       C = volume of sample extract, ml
       D = final volume of extract after concentration (if necessary),
           ml
       E = peak height (or area) of standard response
       F = volume of extract injected into GC to produce  response  B, yl
       H = volume of extract removed for concentration (if necessary), ml
       g = wet weight of sediment initially extracted, g
     % S = sediment percent solids as a decimal fraction
                                  3-335

-------
                              References
1.  American Public Health Association.   Standard  Methods  for the
    Examination of Water and Wastewater  Including  Bottom Sediments
    and Sludges. lUth Edition.   APHA;  New York,  New York.  1193 p.
    (1976).
2.  Environment Canada.   "Analytical Methods Manual."   Inland Waters
    Directorate, Water Quality  Branch; Ottawa, Ontario, Canada (197U).
3.  Forbes, M.  A., Cochrane, W.  P.,  and  Greenhalgh,  R.  "Confirmatory
    Identification of Organophosphorous  Insecticides with  Various
    Reducing Agents."  CIC-CCIW Symposium on Water Parameters,
    19-21 November 1973; Burlington, Ontario, Canada (1973).
h.  Coburn, J., and Chau, A. S.  Y.   "Confirmation  of Pesticide-Residue
    Identity. 8. Organophosphorus Pesticides."   J.  Ass. Offic. Anal.
    Chem. 57:1272-1278 (I9lh).
5-  Anlinker, R., and Beriger,  E.  "Phosphamidon,  Residue  Reviews,"
    Gunther, F. A. (Ed.), 37:l-lU (1971).
6.  Walton, A.   "Methods for Sampling  and Analysis of Marine Sediments
    and Dredged Materials."  Ocean  Dumping Report  1.  Department of
    Fisheries and Environment;  Ottawa, Ontario,  Canada.  7^ p. (1978).
                                 3-336

-------
                   POLYNUCIiEAR AROMATIC HYDROCARBONS
            The basic common structure of this family of compounds is a
fused aromatic ring.  The source of these compounds is diverse as  there
                                         i*
are both natural and man-derived sources.    However, the concern  about
these chemicals results from the fact that many of them are potentially
carcinogenic.
            Polynuclear aromatic hydrocarbons (PAH) are separated  from
the original sample by extraction with dichloromethane, methanol,  or
ethanol.  The extracts are then purified by solvent partitioning.   The
purified extracts are quantified using gas chromatography or fluorimetry.
                      Sample Handling and Storage

            Samples for polynuclear aromatic  hydrocarbon  (PAH)  analysis
should be handled and stored in glass or stainless steel  containers.
During the collection phase, care should be taken not to  contaminate
the sample with lubricants or other hydrocarbon products  as  they may
contain PAH compounds.
            Of the three methods of storing sediment samples, wet,  dry,
or frozen, no evidence was found that any one method is advantageous
or disadvantageous to use.  Similarly, appropriate storage times have
yet to be defined.  In the absence of definitive data, it is recommended
that the procedure for chlorinated hydrocarbon pesticides be followed
and PAH samples be extracted as soon as practical (Figure 3-

             Procedure for Water Samples (Wl. W2,  S1A)2>3

Method 1:  Dichloromethane Extraction/Gas Chromatography
Apparatus
Ultrasonic homogenizer
Filtering apparatus
Vacuum pump
Separatory funnels
* References can be found on page 3-3^7.
                                  3-337

-------
Figure 3-U2  Handling and storage of samples  for polynuclear aromatic hydrocarbon analysis

CORE SAMPLE
,f

JATER SAMPLE DREDGE SAMPLE CORE SECTION
i * *
* * * i*i
FILTER N0 ^)ENT STORE WET FREEZE
1 	 , * i
STORE ' STORE STORE
1 1
EXTRACT EXTRACT

,
+ * 1 	


ill i 1
ANALYZE ANALYZE ANALYZE ANALYZE ANALYZE
LO fv/n fU2) (S1A) (SID) . (S3)
U)
OO
SAMPLE DESIGNATION
PURPOSE
CONTAINER
SAMPLE TREATMENT
PRESERVATIVE
STORAGE TIME
DIGESTION SOLUTION
SAMPLE VOLUME OR WEIGHT
Wl W2 W3 S1A SID S3
Total Water Soluble Used In Mobile Total Total
Cone. Water Elutriate Cone. Sediment Sediment
Cone. Cone. Cone.
G G G G G G
None Filter None None None Freeze
4C 4C None 4C >iC None
(Minimize Air Contact. Keep Field Moist.)
Extract as soon
as possible.
>1 liter >1 liter >1 liter 25-100 g 25-100 g

-------
Vigreux distillation apparatus
Gas chromatography:  Hewlett-Packard 7620 or equivalent.   Balanced dual
      columns (3-5 m by 50 mm O.D. ,  stainless steel columns),  packed vith
      3 percent OV-T on 60/80 mesh acid washed and DCMS treated Gas
      Chrom Q.  Use a nitrogen carrier (35 ml/min) and dual FID detec-
      tors.  Maintain a temperature of 26oC for 8 min and then increase
      the temperature to 300 C at a rate of 8/min
Reagents
Dichloromethane.
Methanol.
Cyclohexane.
Dimethyl Sulfoxide:  when GC analysis is used,  it may be necessary to
      purify commercially available DMSO.  Add 200 ml DMSO to  200 ml
      glass distilled water.  Extract once with 400 ml redistilled iso-
      octane and discard the iso-octane.  Using a rotary evaporator at
      90 to 95C, reduce the solvent .volume until 225 ml of liquid
      (mostly water) has been collected in the condenser  trap.  The
      remaining DMSO is ready for use.
Procedure
            Add 300 ml dichloromethane to 5 & of water sample.  Mix the
sample for 5 niin with an ultrasonic  homogenizer.  The instrument should
be set at approximately 80 percent of full scale.
            A second 5-& water sample should be spiked with 0.1 to
0.2 ml of a dichloromethane solution of standard PAH.  This sample
should be carried through the analytical procedure as an indicator of
extraction efficiency.
            Stopper the mixed sample and allow to sit overnight in the
absence of light.  Decant as much of the aqueous phase as possible.
Filter the remaining liquid under vacuum through a glass  fiber filter.
Wash the filter with a dichloromethane solution and add to the filtrate.
Transfer to a separatory funnel and drain off the dichloromethane
layer.
            Transfer the sample to a Vigreux distillation apparatus and
reduce the sample volume.  Dissolve the sample in 100 ml  ^:1 methanol-
distilled water.
            Extract the PAH into 30 ml cyclohexane.  Add  15 ml dimethyl-
sulfoxide (DMSO) to the cyclohexane extract and shake for 3 min.   Extract
the cyclohexane layer with two more 15-ml portions of DMSO.
            Combine the DMSO extracts and dilute with 90  ml distilled
                                 3-339

-------
water.  Add 25 ml cyclohexane,  shake for 5 min,  and draw off the cyclo-
hexane layer.  Extract the DMSO solution with a  second 25-ml portion of
cyclohexane.  Shake the sample for 5 min during  the extraction process.
            Combine the cyclohexane extracts and transfer the sample to
a Vigreux distillation apparatus.   Evaporate the sample to near dryness.
Dilute the sample to a convenient  volume and analyze "by gas chromatogra-
Phy-
NOTE:  The initial dichloromethane extract may be analyzed directly by
       GC.  However, the DMSO modification is an efficient purification
       procedure (90 to 100 percent.recovery for most PAH).
Calculations
            The PAH concentration can be calculated as follows:
where
     A = weight of standard PAH injected,  yg
     B = observed peak height or area of sample
     C = final sample extract volume, ml
     D = observed peak .height or area produced by standard A
     E = volume of sample injected, ml
     F = volume of sample initially extracted,  
                                 3-3^0

-------
             Procedures for Sediment Samples (SID, 52, S3)

Method 1:  Methanol Extract!on/UV Analysis'*
Apparatus
Soxhlet extraction apparatus
Sodium sulfate column:  extract anhydrous sodium sulfate with 1:1
      benzene-methanol and dry at 12CPC.  Pack in appropriate size
      column
Roto-evaporator
Column packed with copper turnings
Sephadex LH-20 column:  condition 20 g Sephadex LH-20 in 1:1 benzene-
      methanol.  Wash in the same solvent and pack in a glass column,
      1.6 cm I.D., 38 cm height
Alumina-Silica gel column:  activate silica gel and alumina for 2h hr
      at 21CPC.  Deactivate with 3 percent water.  Pack a 1.2 cm I.D.
      glass column with k- ml deactivated silica gel and k ml deacti-
      vated alumina
Silica gel column:  activate silica gel at 210C for 2^ hr.  Slurry in
      n-pentane and pack 12 ml in a 1.2 cm I.D. glass column.
Alumina column:  activate alumina at 210C for 2^ hr.  Deactivate with
      1 peraent water.  Pack 5 ml in a 0.6 cm I.D.  glass column
Hamilton syringes
Elec trobalanc e
Recording UV spectrophotometer with 10 cm quartz cells
Reagents
All reagents are analytical grade.  Solvents should be redistilled when
      necessary.
Methanol.
Benzene.
n-Pentane.
Anhydrous sodium sulfate.
Silica gel.
Alumina.
Sephadex LH-20.
Standard PAH compounds.
2,h ,7-trinitro-9-fluorenone.
Procedure
            Blend the sediment sample.  Weigh out 100 to 150 g wet
                                  3-3^1

-------
weight equivalent of the sediment and transfer to a Soxhlet extraction
thimble.  Place 275 ml methanol and the sample in a Soxhlet extraction
apparatus and extract for 2k hr.   Add 75 iftl benzene and extract for  an
additional 2k hr.
            Transfer the extract to a separatory funnel and partition
the PAH's into n-pentane with three 75-ml aliquots of the solvent.   Wash
the combined n-pentane extract with two 250-ml rinses of distilled water.
Dry the extract by passing through an anhydrous sodium sulfate column
and collect in a round-bottomed flask.   Concentrate the n-pentane extract
to approximately 1 ml on a roto-evaporator at room temperature.
            Transfer the sample to a column of copper turnings to
remove elemental sulfur.  Elute the sample from the column using
benzene-pentane (l:l) as an eluent.  Reduce the eluate volume  to
approximately 1 ml.
            Place the sample on the Sephadex LH-20 column and  elute
with benzene-methanol (l:l) at a flow rate of 6 ml/min.  Discard the
first 50 ml of eluate.  Collect the second 50 ml and evaporate to
dryness in a rotary evaporator at room temperature.
NOTE:  The column can be reused by washing with 100 ml of solvent
       (.1:1 benzene-methanol) and repacking the column as necessary
       to maintain the flow rate.
            Take up the aromatic  residue in 1 ml n-pentane and transfer
to an alumina-silica gel column.   Elute the column with 20 ml  n-pentane
and discard.  Rinse the rotary evaporation flask with 2 ml methylene
chloride and add to the alumina-silica gel column.   Elute the  column
with an additional 13 ml methylene chloride and retain the eluate in
a 100-ml round-bottomed flask.
            Add 20 mg Trinitrofluorenone (TNF) to the sample and evapo-
rate to complete dryness.   Wash the sample 5 times with 2 ml rinses  of
n-pentane.  Withdraw each wash through a cotton pad using a pipet and
discard.
            Dissolve any excess TKF and the PAH complexes in methylene
chloride and transfer to an activated silica gel column.   Elute the
PAH complexes with 75 ml methylene chloride and evaporate to near
dryness (0.5 - 1.0 ml).
            Transfer a .small aliquot (< 100 yl)  to the aluminum pan
                                 3-3^2

-------
of an electrebalance, air dry, and weigh.
            Dry the remainder of the PAH concentrate under a stream of
nitrogen at room temperature.  Dissolve the residue in 0.5 nil l:k methy-
lene chloride:pentane and apply to an alumina column.  This solution
should not contain more than 200 yg of material.   Elute the sample with
pentane containing an increasing percentage of methylene chloride; 95 nil
h percent methylene chloride, 70 ml 15 percent methylene chloride, 30 ml
20 percent methylene chloride, 30 ml 30 percent methylene chloride, and
20 ml 100 percent methylene chloride.
            Collect the eluate in seven fractions based on general ring
types.  The volume of each fraction and the compounds in each fraction
are as follows:
      Fraction No.  Volume, ml     Compounds   Wavelength, nm
1
2
3
20
25
60
u
5
6
7
20
35
35
25
                                PCB's
                                Phenanthrene
                                Anthracene
                                Pyrene
                                Fluoranthene
                                Chrysene
                                Benzanthracene
                                Benzopyrenes
                                Perylene
                                Benzoperylene
                                Anthanthrene
                                Coronene
293
252
333
286
26?
287
383
382
U28
338
            Adjust the final volume of each fraction to 25 ml by  eva-
poration or dilution with methylene chloride.   Determine the spectra
of each fraction with a recording UV spectrophotometer  using a 10-cm
quartz cell.  Compare the sample spectra to standard PAH spectra  and
quantify each PAH at the specific wavelength listed  above.
Calculations
follows:
            Calculate the concentration of each PAH compound as
               PAH yg/kg (wet weight)  = (x)(0.025)ClOOO)
                         (dry elght)  .
where
       X = PAH concentration in extract,  ug/
                                 3-3^3

-------
       0.025 = final volume of fraction extract, I
           g = wet weight of sediment sample extracted, g
         % S = sediment percent solids (expressed as a decimal fraction)

Method 2:  Ethanol Extraction/UV Spectrophotometry5
Apparatus
Erlenmeyer flasks
Separatory funnels
Florisil column:   add 30 g florisil to a UO- by ^00-mm glass column
      fitted with a coarse fitted glass disc.  Cover with 60 g WaaSOii
Roto-evaporator
Thin-layer chromatography plates and development tanks
Fine fitted Buchner funnel
Recording spectrophotofluorimeter, Aminco-Bowman or equivalent
Reagents
All reagents should be of analytical grade.
Ethanol.
Potassium hydroxide.
Boiling stones.
Glass wool.
Iso-octane.
Benzene.
Dimethyl sulfoxide:  if samples are to be quantified by gas chroma-
      tography, purify the DMSO as follows:
      Add 200 ml DMSO to 200 ml glass distilled water.  Extract
      once with UOO ml redistilled iso-octane.   Discard the iso-
      octane.  Using a rotary evaporator at 90 to 95C, reduce the
      solvent volume until 225 ml of liquid (mostly water) has been
      collected in the condenser trap.  The remaining DMSO does not
      have to be dried before use.
Na2SOlt.
Toluene.
Hexadecane.
PAH standards.
Procedure
            Blend a sediment sample and weigh out a 10- to 20-g wet
weight equivalent of the sample.  Transfer to a flask and add 100 ml

                                3-3^1*

-------
ethanol, 5 g KOH, and a few boiling stones.   Reflux the sample for
1.5 hr.
            Pour the suspension into a 250-ml Erlenmeyer flask and
allow the sediment to settle by gravity for  5 min.   Decant the alcohol
through a glass wool plug into a l- separatory funnel containing
150 ml distilled water.
            Wash the sediment twice with 50-ml portions of ethanol.
Filter each wash through the glass wool plug and add to the separatory
funnel .
            Extract the water/ethanol mixture three times with 200 ml
iso-octane.  Combine the iso-octane extracts and wash four times  with
200-ml portions of warm (60C) water.
            Wash a Florisil column with 100  ml of iso-octane.   Place
the iso-octane extract on the column and allow the  solvent to  drain.
Wash the column with two 100-ml portions of  fresh iso-octane and
allow the column to drain briefly between each addition.
            Elute the PAH's from the column  with three 100-ml  portions
of benzene.  Collect the eluate in a round-bottomed flask and  reduce to
5 ml on a rotary evaporator.  Add 50 ml iso-octane  and reduce  the
volume to 5 ml, again on a rotary evaporator.
            Extract the iso-octane extract with three 5-ml portions
of dimethyl sulfoxide (DMSO) .  Combine the DMSO extracts with  30  ml
distilled water.*  Extract the PAH back into iso-octane with two  10-ml
portions of the solvent. -Combine the iso-octane extracts and  wash
three times with 20 ml distilled water.   Dry the sample by passage
through 10 g NaaSOi* in a 15-ml coarse fritted glass Buchner funnel.
*  The following procedure can be used if the PAH are  to be  quantified
  using gas chromatography:
        Add the DMSO extract to U5 ml distilled  water.  Extract the
        aqueous solution with two 12. 5-ml portions of  cyclohexane.
        Shake the mixture 5 min each time and draw off the cyclohexane
        layer.  Combine the extracts and transfer to a 15 -cm Vigreux
        column.  Evaporate to near dryness,  transfer to a pear-shaped
        flask.  Rinse the Vigreux flask with cyclohexane and add the
        rinsing to the pear-shaped flask.  Continue evaporating the
        sample to a final volume of approximately 20 yl.
        Analyze using a gas chromatograph .  Use  the instrument opera-
        ting conditions specified for water  samples.
                                3-3^5

-------
            Reduce the sample size to approximately 0.1 ml using rotary
evaporation and a stream of nitrogen.  Spot the sample and a 10-ng PAH
standard on a cellulose-acetate thin-layer plate.   Support the plate
approximately 5 cm from the bottom of a development tank with the
plate extending approximately 2 cm above the tank.   Add sufficient
developing solution (ethanol:toluene:water, 17:^:^0 to wet the "bottom
of the plate.  Partially cover the top of the development tank.
            Locate the PAH band with long wave ultra-violet light.
The PAH band should be the lowest band on the plate with an approxi-
mate Rf of 0.3 after 2 hr of development.
            Scrape off the PAH band while still wet and place in a
15-ml fine fritted glass Buchner funnel.  Wash the  sample four times
with k-ml- portions of hot (65C) methanol.  Use gentle suction to
separate the solvent.   Add the combined methanol extract to 10 ml of
l:k hexadecaneiso-octane.  Reduce the sample volume to 2 ml on a
roto-evaporator.
            Exite the samples and standards, containing no more than
200 mg PAH, at 365 nm with a spectrophotofluorimeter.   Record the
sample spectrum from 375 to 500 nm.  Quantify the PAH concentration
based on the maxima at ^30 nm relative to an artificial baseline
between Itl8 and UkQ nm.
Calculations
            Prepare a standard curve of ng PAH vs.  sample fluorescence.
Calculate the weight of PAH in the sample extracts:
                 PAH yg/kg (wet weight) = (X)(V)(l000)

                 PAH Wg/kg (dry .eight) =
where
       X = PAH concentration in final extract, yg/&
       V = volume of final extract, 5L
       g = wet weight of sediment extracted, g
     % S = sediment percent solids (expressed as a  decimal fraction)
                                3-3U6

-------
                          References
Blumer, M., and Youngblood, W. W.   "Polycyclic Aromatic  Hydrocarbons
in Soils and Recent Sediments."  Science 188:53-55 (1975).
Acheson, M. A., Harrison, R. M., Perry, R.,  and Wellings, R.  A.
"Factors Affecting the Extraction and Analysis of Polynuclear
Aromatic Hydrocarbons in Water."  Water Research 10:207-212 (1976).
Hoffman, D., and Wynder, E. L.  "Short Term Determination of
Carcinogenic Aromatic Hydrocarbons."  Anal.  Chem. 32:295-296 (1960).
Giger, W., and Blumer, M.  "Polycyclic Aromatic Hydrocarbons in
the Environment:  Isolation and Characterization by Chromatography,
Visible, Ultraviolet, and Mass Spectrometry."  Anal.  Chem.  ^
1171 (1971*).
Dunn, B. P.  "Techniques for Determination of Benzo(a) Pyrene in
Marine Organisms and Sediments."  Environ. Sci. and Tech. 10:
1018-1021 (1976).
                            3-3^7

-------
                               PHENOLICS

            Phenols are hydroxy-derivatives of benzene and related
compounds.  The colorimetric procedures used to quantify phenols
are not specific for one phenolic compound, but to the general phenol
structure.  Because of the difficulty in preparing standard phenol
mixtures for all samples, phenolic compounds are reported as an equi-
valent amount of phenol.

                      Sample Handling and Storage

            Phenol samples should be stored in glass containers.
When immediate analysis is not possible, samples may be preserved with
the addition of 1 g/ copper sulfate and acidification with phosphoric
acid to pH < U.  The samples should also be maintained at UC until
analysis.  However, even with the use of these preservatives, samples
should be analyzed within 2k hr.  This information is summarized in
Figure 3-^3.
            It is recommended that only wet sediment samples be
utilized for phenol analysis.  This cautionary approach is based on
the fact that dried samples may lose phenol as a result of biological
degradation, and both dried and frozen samples may lose phenol as a
result of volatilization during the drying and/or thawing cycles.
Samples should also be processed within the 2^-hr period specified
for water samples.

              Procedures for Water Samples (Wl, W2, S1A)
                                                             Afc
Method 1:  Distillation, U-aminoantipyrine Colorimetric1'2'3
Apparatus
Distillation apparatus, all glass consisting of a l- pyrex distilling
      apparatus with Graham condenser
pH meter
Spectrophotometer, for use at U6o or 510 nm
* References for this section are on page 3-360.
                                3-3U8

-------
                 Figure  3-U3.      Handling and storage of  samples  for  phenol  analysis


ACIDIFY
1
STORE
1
DISTILL
1
ANALYZE
(Wl)
U)
WATER SAMPLE CORE SAMPLE
t | .
i * 	 *
FILTER N 5tENT DREDGE SAMPLf CORE SECTION
i
1 i

ACIDIFY STORE WET
i
STORE
1
' '
4 +
DISTILL 1 l ELUTRIATE DISTILL
i 1 i
ANALYZE ANALYZE ANALYZE
(W2) (S1A) (SID)

 hr
                                                            2k hr
                                                                                              2k hr
DIGESTION SOLUTION
                        Distillation   Distillation
                                                          Distillation
                                                                                            DistiIlation
SAMPLE VOLUME OR WEIGHT
                          500 ml
                                       500 ml
                                                             500 ml
                                                                                              10-50 g

-------
Funnels
Filter paper
Membrane filters
Separatory funnels, 500 or 1QOO ml
Nessler tubes, short or long
Reagents
Phosphoric acid solution, 1 + 9:  dilute 10 ml of 85 percent H.3POi, to
      100 ml with distilled water.
Copper sulfate solution:  dissolve 100 g CuSOi^   5^2 0 in distilled
      water and dilute to 1 .
Buffer solution:  dissolve 16.9 g NIUC1 in 1^3 ml cone. NHi,OH and
      dilute to 250 ml with distilled water.  Two ml should adjust
      100 ml of distillate to pH 10.
Aminoantipyrine solution:  dissolve 2 g of k AAP in distilled water
      and dilute to 100 ml.
Potassium ferricyanide solution:  dissolve 8 g of KsFeCciOe  in
      distilled water and dilute to 100 ml.
Stock phenol solution:  dissolve 1.0 g phenol in freshly boiled and
      cooled distilled water and dilute to 1  .   1 ml = 1 mg phenol.
Working solution A:  dilute 10 ml stock phenol solution to 1  with
      distilled water.  1 ml = 10 ug phenol.
Working solution B:  dilute 100 ml of working solution A to 1000 ml
      with distilled water.  1 ml = 1 ug phenol.
Chloroform .
Procedure
            The first step in the procedure is a distillation to
isolate phenolic compounds from possible interferring substances in
the sample.  Add sufficient 1+9 phosphoric acid to a 500-ml water
sample (Wl, W2, SLA.) to lower the pH to approximately k.  Add 5 ml
copper sulfate solution to the sample.
NOTE 1:  Omit the addition of H3P04 and CuSOi* to the sample if these
         reagents have previously been added to  the sample as pre-
         servatives.
NOTE 2:  The addition of HsPOit and CuSOii serves the dual purposes of
         inhibiting the biological degradation of phenol and
         removing the interference due to sulfur compounds.
            Transfer the acidified sample to the distillation
apparatus.   Distill the sample and collect the distillate.   When
U50 ml of distillate have been collected, temporarily stop  the

                                3-350

-------
distillation process.  After boiling has ceased in the distillation
flask, add 50 ml warm distilled water to the flask.  Resume sample
distillation until a total of 500 ml distillate has been collected.
            If the distillate is turbid, filter the sample through a
O.H5-y pore-sized membrane filter.  Analyze the sample using either
direct colorimetry (Method 1A) or chloroform extraction (Method IB).
The direct method is for samples with, phenol concentrations in the
range of kQ to 1000 yg/ and the extraction technique is for samples
with phenol concentrations in the range of 5 to 50 jjg/fc.
            Method 1A:  Direct Colorimetry.  Prepare the following
set of phenol standards in 100-ml volumetric flasks by pipetting the
indicated volume of phenol working solution A.
                                                       Phenol
      ml of working solution A                       cone. , yg/
                0                                         0.0
                0.5                                      50.0
                1.0                                     100.0
                2.0                                     200.0
                5.0                                     500.0
                8.0                                     800.0
               10.0                                    1000.0
            Pipet either 100 ml of standard, 100 ml of distillate,
and/or an aliquot of the sample diluted to 100 ml into an Erlenmeyer
flask.  Add 2 ml of ammonia buffer solution and mix.   The resultant
pH should be 10 +_ 0.02.
            Add 2.0 ml aminoantipyrine solution and mix.  Add 2.0 ml
potassium ferricyanide solution and mix.  Allow 15 min for color
development and measure the absorbance of the sample at 510 nm
relative to a reagent blank.
            Method IB:  Chloroform Extraction.  Low concentrations
of phenol can be concentrated by chloroform extraction to enhance
detection.   The phenol-aminoantipyrine color complex is developed
as in Method 1A and concentrated by chloroform extraction.
            Prepare a series of phenol standards by pipetting the
indicated volume of phenol working solution B into a series of
separatory funnels and diluting each to 500 ml with distilled
water.
                                3-351

-------
                                                       Phenol
      ml of working solution B                       cone.,  yg/&
                 0.0                                     0.0
                 3.0                                     6.0
                 5.0                                    10.0
                10.0                                    20.0
                20.0                                    UO.O
                25.0                                    50.0
            Place 500 ml of sample distillate or an aliquot  diluted
to 500 ml in a separatory funnel.  The distillate should "be  prepared
as described in Method 1A and should not contain more than 25 yg
phenol.  Add 10 ml ammonia buffer solution and mix.  The pH  of the
sample should be 10 +_ 0.02.
            Add 3.0 ml aminoantipyrine solution and mix.  Add 3.0 ml
potassium ferricyanide solution and mix.
            After 3 min, extract with 25 ml of chloroform.   Shake
the separatory funnel at least 10 times, let CHCla settle,  shake
again 10 times, and let chloroform settle again.
            Filter the chloroform extracts through filter paper.
Do not add any chloroform to compensate for any chloroform that may
be lost during the filtration process.
            Measure the absorbance of the standards and samples at
k6Q run relative to a reagent blank.
Calculations
            Prepare a standard curve by plotting the absorbance
value of standards vs. the corresponding phenol concentrations.
Obtain concentration value of sample directly from standard  curve.

Method 2:  Distillation, MBTH Colorimetric1
            Phenolic compounds are separated from the sample matrix
by distillation.  The phenols are coupled with 3-methyl-2 benzo-
thiazolinone hydrazone hydrochloride (MBTH) in an acidic solution.
The complex is then oxidized with eerie ammonium sulfate to  produce
a color proportional to the original phenolic concentration.
Apparatus
Distillation apparatus:  all glass consisting of a 1-& pyrex
      distilling apparatus with Graham condenser
                                3-352

-------
pH meter
Spectrophotometer

Funnels
Filter paper

Membrane filters

Separator/ funnels

Reagents

Copper sulfate solution:  dissolve 100 g CuSOi*   5H20 in distilled
      vater and dilute to 1 .
Sulfuric acid, 1 H_:  add 28 ml of cone. H2SOi, to  900 ml of distilled
      water, mix, and dilute to 1 .

MBTH solution, 0.05 percent:  dissolve 0.1 g of 3-methyl-2-benzothia-
      zolinone hydrazone hydrochloride in 200 ml  of distilled vater.

Ceric ammonium sulfate solution:  add 2.0 g of Ce(SOit)2 *  2(NHit)2SOit 
      2H20 and 2.0 ml of cone. H2S04 to 150 ml of distilled water.
      After the solid has dissolved, dilute to 200 ml with distilled
      water.
Buffer solution:  dissolve in the following order, 8 g of sodium
      hydroxide, 2 g EDTA (disodium salt), and 8  g "boric acid in
      200 ml of distilled water.  Dilute to 250 ml with distilled
      water.
Working buffer solution:  make a working solution by mixing an
      appropriate volume of buffer solution with  an equal volume
      of ethanol.

Chloroform.

Stock phenol:  dissolve 1.00 g phenol in 500 ml of distilled water
      and dilute to 1000 ml.  Add 1 g CuSOi, and 0.5 ml cone. H2S04
      as a preservative.  1.0 ml = 1.0 mg phenol.
Standard phenol solution A:  dilute 10.0 ml stock phenol solution
      to 1000 ml.  1.0 ml = 0.01 mg phenol.

Standard phenol solution B:  dilute 100.0 ml of standard phenol
      solution A to 1000 ml with distilled water.  1.0 ml = 0.001 mg
      phenol.

Procedure

            Transfer 500 ml of sample (Wl, W2, and/or S1A) to a 1-5,

distillation flask.  Add 5 ml 10 percent copper sulfate solution and

adjust the pH to approximately k with 1 N_ sulfuric acid.

            Distill over approximately ^50 ml of  sample and interrupt

the distillation process.  Add 50 ml warm distilled water to the

distillation flask and resume distillation until  a total of 500 ml

                                3-353

-------
distillate has been collected.   If the distillate is  turbid,  filter
through a prewashed, 0.^5-y pore-sized membrane  filter.
            If the phenol concentration is above 50 yg/&, the concen-
tration can be determined directly (Method 2A).   If the phenol con-
centration is less than 50 yg/, the colored  complex  is concentrated
by solvent extraction prior to  quantification (Method 2B).
            Method 2A:  Direct  Colorimetry.   To  100 ml of standard,
distillate, or sample aliquot diluted to 100  ml, add  h ml MBTH
solution and mix.
            Allow 5 min for the coupling reaction and add 2.5 ml
eerie ammonium sulfate solution.  Mix.
            After a second 5-min period, add  7 ml working buffer
solution.  Wait 15 min for color development  and measure the  sample
absorbance at 520 nm relative to a reagent blank. The color  is
stable for k hr.
            Method 2B:  Solvent Extraction.   Transfer the 500-ml
distillate to a 1-A separatory  funnel and add k  ml MBTH solution.
Mix and wait 5 min.
            After 5 min, add 2.5 ml eerie ammonium sulfate  solution.
Wait 5 min and add 7 ml of working buffer solution.
            Allow 15 min for color developments  and add 25  ml chloro-
form.  Shake the separatory funnel at least 20 times. Drain  the
chloroform layer through filter paper.
            Measure the absorbance of the sample at ^90 nm  relative
to a reagent blank.
Calculations
            Prepare a standard  curve by plotting the  absorbance of
the phenol standards against known concentrations. Compare sample
absorbance measurements to the  standard curve to determine  phenol
concentrations in the samples.

-------
                 Procedures for Sediment Samples (SID)

            The phenol procedure for sediments is essentially the same
as that used for water samples.  Phenols are separated from the sample
matrix and possible interferences by distillation.   Phenols in the
distillate are complexed with either l*-aminoantipyrine or 3-methyl-2-
benzothiazolinone hydrazone hydrochloride (MBTH) and measured colori-
metrically .
Method 1:  Distillation, U-aminoantipyrine Colorimetric
Apparatus
Distillation apparatus, all glass, with l- distillation flask
Separatory funnels, 1 , with Teflon stopcocks
Spectrophotometer, with U60-nm filter
Reagents
Phenol stock solution:  dissolve 1.000 g phenol in distilled water
      and dilute to 1 H with distilled water.  1.0 ml = 1.0 mg phenol.
Phenol working solution:  pipet '20.0 ml phenol stock solution into a
      1-Jl volumetric flask and dilute to volume with distilled water.
      1.0 ml = 20 ug phenol.  Prepare daily.
Phenol standard solution:  pipet 25.0 ml phenol working solution into
      a 500-ml volumetric flask and dilute to volume with distilled
      water.  1.0 ml = 1.0 ug phenol.  Prepare daily.
Ammonium chloride solution:  dissolve 67. 5 g WHitCl in 570 ml cone.
            and dilute to 1  with distilled water.
Aminoantipyrine solution:  dissolve 2.0 g U-aminoantipyrine in
      distilled water and dilute to 100 ml.
Potassium ferricyanide solution:  dissolve 8.0 g KsFeCciOs  in distilled
      water and dilute to 100 ml.
Chloroform, reagent grade.
Phosphoric acid solution:  mix 10 ml phosphoric acid with distilled
      water and dilute to 100 ml.
Copper sulfate solution:  dissolve 10 g CuSOi*  5^0 in distilled water
      and dilute to 100 ml.
Procedure
            Place an aliquot of wet sediment (SID),  10  to 50  g,  con-
taining not more than 50 yg phenol, in a l- distillation flask.  Add
550 ml distilled water.
                                3-355

-------
            Add 5 ml 10 percent copper sulfate solution, 5 ml phosphoric
acid solution, and a. few drops of methyl orange indicator.
NOTE:  The addition of CuSOn and HsPOi* can be omitted if the sample was
       previously preserved.
            Add a few boiling stones and distill 500 ml of distillate.
NOTE:  If oil is present in the distillate, filter the sample through
       two thicknesses of dry No. 12 Whatman filter paper to remove the
       oil.  Collect the filtered sample in a l- separatory funnel.
            The phenol concentration can be quantitated using direct
colorimetry if the concentration is above 50 VgA (Method 1A) or by
solvent extraction if the concentration is less than 50 yg/ (Method
IB).
            Method 1A:  Direct Colorimetry.  Pipet 100 ml distillate
into an Erlenmeyer flask and add 2 ml ammonia buffer.  Mix.  Sample
pH should be 10 +_ 0.02.
            Add 2 ml aminoantipyrine and. mix.  Add 2 ml potassium
ferricyanide and mix again.  Allow 15 min for color development.
Measure the absorbance of the sample at 510 nm relative to a reagent
blank.
            Method IB:  Solvent Extraction.  Transfer 500 ml distillate
to a 1-& separatory funnel.  Add 3 ml ammonia chloride solution and
mix.  Add 3 ml aminoantipryrine solution and mix.   Add 3 ml potassium ferri-
cyanide and mix again.  Allow color to develop 3 to 5 min.
            Add 25 ml chloroform and shake vigorously for 30 sec.
Allow layers to separate and shake again for 30 sec.
            Draw off the chloroform layer through filter paper or a
cotton pledget.  Measure the absorbance of the sample at k6o nm
relative to a reagent blank.
Calculations
            Prepare a standard curve based on the absorbance of the
standard phenol solutions.  Determine the phenol concentrations in
the distillate by comparing sample absorbance with the standard
curve.
                    i   h   i  *.   -  v.^   (A)(B)(IOOO)
               Phenol ug/kg (wet weight) = L^	
                                                 o
                                3-356

-------
                        /i   i A       ^\   U)(B)ClOOO)
               Phenol yg/kg (dry weight) =   |\^ ^

where
       A = phenol concentration in distillate, yg/

       B = total volume of distillate,  (0.5 & as written)

       g = wet weight of sediment, g

     % S = percent solids in sediment (expressed as a decimal fraction)


Method 2:  MBTH Color imetric

Apparatus
Distillation apparatus, all glass, with l- distillation flask

Separatory funnels, 1 , with Teflon stopcocks

Spectrophotometer , with ii90-nm filter

Reagents

Copper sulfate solution:  dissolve 100 g CuSOi*   5HzO in distilled
      water and dilute to 1 A.
Sulfuric acid, 1 N_:  add 28 ml of cone.  HaSOi* to 900 ml of distilled
      water, mix, and dilute to 1 .

MBTH solution, 0.05 percent:  dissolve 0.1 g of 3-m ethyl- 2-benzo-
      thiazolinone hydrazone hydrochloride in 200 ml of distilled
      water.
Ceric ammonium sulfate solution:  add 2.0 g of Ce( SOOa *
      SO*  2H20 and 2.0 ml of cone. H2SO. to 150 ml of distilled
      water.  After the solid has dissolved, dilute to 200 ml with
      distilled water.
Buffer solution:  dissolve in the following order:  8 g of sodium
      hydroxide, 2 g EDTA (disodium salt) and 8 g boric acid in
      200 ml of distilled water.  Dilute to 250 ml with distilled
      water .

Working buffer solution:  make a working solution by mixing an
      appropriate volume of buffer solution with an equal volume of
      ethanol . -
Chloroform.

Stock phenol:  dissolve 1.00 g phenol in 500 ml of distilled water
      and dilute to 1 i.  Add 1 g CuS04 and 0.5 ml cone. HaSOi* as
      preservative.  1.0 ml = 1.0 mg phenol.
Standard phenol solution A:  dilute 10.0 ml of stock phenol solution
      to 1 H.  1.0 ml = 0.01 mg phenol.
Standard phenol solution B:  dilute 100 ml of standard phenol solution
      A to 1  with distilled water.  1.0 ml = 0.001 mg phenol.
                                3-357

-------
Procedure
            Weigh out a 10- to 50-g aliquot of blended,  wet sediment  and
transfer to a l- distillation flask.   The sediment sample should not
contain more than 50 yg phenol.  Add 550 ml distilled water.
            Add 5 ml 10 percent copper sulfate solution  and 5 ml
phosphoric acid solution.  The pH of the sediment suspension should be
approximately h.
NOTE:  This step can be omitted if the sample was previously preserved.
            Add a few boiling stones and distill over 500 ml of sample.
The phenol concentration can be quantified using direct  colorimetry if
the concentration is above 50 yg/ (Method 2A) or by solvent extraction
if the concentration is less than 50 yg/& (Method 2B) .
            Method 2A:  Direct Colorimetry.  To 100 ml of distillate
or a smaller aliquot diluted to 100 ml, add k ml MBTH solution and mix.
            Allow 5 min for the MBTH-phenol coupling reaction and
add 2.5 ml eerie ammonium sulfate solution.  Mix the solution.
            Five minutes later, add 7  ml working buffer  solution.   Wait
15 min for color development and measure the sample absorbance at
520 nm relative to a reagent blank.  The color is stable for h hr.
            Method 2B:  Solvent Extraction.  Transfer the 500-ml
distillate to a l- separatory funnel.  Add k ml MBTH solution, mix,
and allow 5 min for the phenol-MBTH coupling reaction.
            Add 2.5 ml eerie ammonium sulfate and again  wait 5 min.
Add 7 ml working buffer solution and allow 15 min for color development.
            Add 25 ml chloroform to the separatory funnel and shake
vigorously for 30 sec.  Let the layers separate and again shake for
30 sec.  Drain the chloroform layer through filter paper.  Measure
the absorbance of the sample at ^90 nm relative to a reagent blank.
Calculations
            Prepare a standard curve by plotting the absorbance of the
phenol standards against known concentrations.  Compare  sample
absorbance measurements to the standard curve to determine phenol
concentrations in the distillates.  Calculate the sediment phenol
concentrations :
               Phenol yg/kg (wet weight) =
                                3-358

-------
              Phenol yg/kg (dry weight)  =
where
       A = distillate phenol concentration,
       B = volume of distillate, H
       g = wet weight of sediment, g
     % S = percent solids in sediment (expressed as a decimal fraction)
                                3-359

-------
                              References
1.  Environmental Protection Agency.   "Methods for Chemical Analysis  of
    Water and Wastes."  Environmental Monitoring and Support Laboratory,
    EPA; Cincinnati, Ohio (1979).
2.  American Public Health Association.   Standard Methods for the
    Examination of Water and Wastewater  Including Bottom Sediments and
    Sludges. iVbh Edition.  APHA;  New York,  New York.  1193 p. (1976).
3.  American Society for Testing Materials.   Book of ASTM Standards,..
    Part 31. Water.  American Society for Testing Materials; Philadelphia,
    Pennsylvania (1976).

U.  U. S. Department of  the Interior.  "Chemistry Laboratory Manual for
    Bottom Sediments."  Great Lakes Region Committee on Analytical
    Methods, U. S. Department of the  Interior; Chicago, Illinois.
    96 p. (1968).
                                 3-360

-------
MISCELLANEOUS ANALYSIS

     Chlorine Demand
     Biochemical Oxygen Demand
     Chemical Oxygen Demand
     Sediment Oxygen Demand
                                 3-361

-------
                            CHLORINE DEMAND

            The chlorine demand of a sample is the difference between
 the  amount of chlorine applied and the amount of free, combined, or
 total  available chlorine remaining at the end of the contact period.
 The  demand is caused by substances that can be oxidized by, or react
 with,  chlorine.  Examples of specific compounds that will be included
 in the chlorine demand are reduced inorganic ions such as ferrous iron,
 manganous manganese, nitrite, sulfide and sulfite, ammonia and amino
                                                 i*
 compounds, and aromatic compounds such as phenol.
            The original purpose of a chlorine demand test was to
 determine the amount of chlorine that had to be added to a water source
 to achieve a free chlorine residual for disinfection purposes.  This
 was  accomplished by adding a known quantity of chlorine to the sample
 and  observing residual chlorine concentrations over time.  If residual
 chlorine goes to zero during the test, the test should be terminated
 at that point in time or rerun at a higher initial chlorine concentra-
 tion.  Since the chlorine demand will vary with the initial chlorine
 dose,  pH, temperature, and time of contact, all test conditions should
 be recorded.

                      Sample Handling and Storage

            Many of the compounds that contribute to the chlorine
 demand of a sample can also be oxidized by oxygen.   Therefore, sample
 processing should begin as soon as possible after collection using
 either total water samples (Wl), filtered water samples (W2, S1A),  or
 field moist sediment samples (SID).   The processes  of air drying and
 freezing/thawing can alter chlorine demand of the samples and are not
 recommended.   This information is summarized in Figure 3-hh.
* References for this section are found  on page 3-372.
                                3-362

-------
OJ

U)
CT\

DREDGE SAMPLE
4
i
P
T
CORE SECTION
1
SAMPLE
DESIGNATION
Wl
W2
W3
S1A
SIB
SIC
SID
S2
S3
           PURPOSE
           CONTAINER
           SAMPLE TREATMENT
           PRESERVATIVE
           STORAGE TIME
           DIGESTION SOLUTION
           SAMPLE VOLUME OR WEIGHT
                                    Total  Water
                                       Cone.
                                       G,P
                                       None
Soluble    Used In
 Water    Elutriate
 Cone.
 G.P
G,P
                                                  Filter      None
            Mobile
           " Cone.
G,P
                                                                          None
  Total
Sediment
  Cone.

   G,P
                                                                                                              None
                                      2l( hr
                                                  24 hr       <1 w
                          (Minimize Air Contact.  Keep Field Moist.)


                        24 hr
                                                                                                              <1 w
                                     250 ml       250 ml
                       250 ml
                                                                                                               2 g
                        Figure 3-hk.      Handling and  storage of samples  for chlorine  demand  analysis

-------
              Procedures for Water Samples (Wl,  W2.  S1A).1
Apparatus
Stirrer and magnetic stirring "bar

500-ml Erlenmeyer flasks

50-ml buret

Reagents
Standard chlorine solution:  prepare by bubbling chlorine gas  through
      distilled water or by diluting household bleach, a hypochlorite
      solution, to a suitable concentration.   The concentration of the
      chlorine solution should be sufficiently strong that the volume
      of the sample will not be increased by  more than 5 percent after
      the addition of the chlorine solution.   The useful life  of the
      solution can be extended by storage in  a dark or brown glass-
      stoppered bottle.  However, a chlorine  solution is unstable
      and must be standardized each time it is used.
      In order to standardize the chlorine solution,  place 2 ml
      glacial acetic acid and 25 ml distilled water in a flask.   Add
      approximately 1 g KI.  Pipet a convenient volume of chlorine
      solution into the flask.  Titrate with  a 0.025 H_ sodium  thio-
      sulfate solution to a pale yellow color.  Add 1 to 2 ml  of
      starch indicator solution and continue  the thiosulfate titration
      to the disappearance of th.e blue color.  A blank consisting of
      2 ml of glacial acetic acid, 25 ml of distilled water, and 1 g
      KI should also be titrated to correct for any chlorine demand
      or residual chlorine in the water or reagents.
      It will be necessary to conduct a blank titration to correct the
      results for reagent impurities such as:  (a) the free iodine or
      iodate in the potassium iodide that liberates extra iodine;  or
      (b) the traces of reducing agents that  might reduce some of the
      iodine liberated.  Take a volume of distilled water corresponding
      to the sample used for titration during the standardization pro-
      cedure, add 2 ml acetic acid, 1 g KI, and 1 ml starch solution.
      Since the blank may be either positive  or negative, it will be
      necessary to perform either blank titration A or B, whichever
      applies:
      a_.  Blank titration A:  if a blue color develops, titrate with
          0.01 N_ or 0.025 N. sodium thiosulfate to the disappearance
          of the color and record results.
      b_.  Blank titration B:  if no blue color occurs, titrate with
          0.0282 N_ iodine solution until a blue color appears.  Back-
          titrate with 0.01 W_ or 0.025 I[ sodium thiosulfate, and
          record the difference as titration  B.
      Before calculating the chlorine consumed, subtract blank
      titration A from the sample titration,  or, if necessary, add the
      net equivalent value of blank titration B.

-------
Glacial acetic acid, CH3COOH.

Potassium iodide crystals, KI .

Standard sodium thiosulfate, 0.1 N_:  dissolve 25 g Na2S20s   5HaO in
      1 H freshly boiled distilled water and standardize the solution
      against potassium biniodate or potassium dichromate after at
      least 2 weeks storage.  Use boiled distilled water and add a few
      milliliters CHCla to minimize bacterial decomposition of the
      thiosulfate solution.

      Standardize the 0.1 N_ sodium thiosulfate titrant using either
      (a) the biniodate method or (b_) the dichromate method:

      a_.  Biniodate method:  dissolve 3.2^9 g primary standard quality
          anhydrous potassium biniodate, 0(103)2, in distilled water
          and dilute to 1 X, to yield a 0.1000 N_ solution.  Store in a
          glass-stoppered bottle.

          To 80 ml distilled water, add, with constant stirring,
          1 ml cone. H2SOi*, 10 ml 0.100 N_ 0(103)2, and 1 g KI.
          Titrate immediately with sodium thiosulfate titrant until
          the yellow color of the liberated iodine is almost  discharged.
          Add 1 ml starch indicator solution and continue the titration
          until the blue color disappears.
      b_.  Dichromate method:  dissolve ^.90^ g primary standard
          quality anhydrous potassium dichromate, K2Cr20?, in distilled
          water and dilute to 1000 ml to yield a 0.100 N_ solution.
          Store in a glass-stoppered bottle.
          Proceed as in the biniodate method, with the following
          exceptions:  substitute 10.00 ml 0.1000 N foCraO? for the
          0(103)2 and let the reaction mixture stand for 6 min in
          the dark before titrating with the Na2S20s titrant.

      The normality of the thiosulfate titrant can be calculated as:

                             Normality
             (Volume oxidizing agent) (Normality oxidizing agent)
                          (Volume Na2S20s consumed)

Dilute sodium thiosulfate titrant, 0.01 If or 0.025 N.:   dilute 100 ml
      (0.01 N) or 250 ml (0.025 N.) of 0.1 N_ sodium thiosulfate to 1 
      with distilled water.   To improve the stability of these titrants,
      the standard thiosulfate should be aged several weeks and the
      distilled water should be fresh.
      Standardize this solution daily in accordance with the directions
      given above, using 0.01 N or 0.025 N KH(lOs)2  or K2Cr207.  (To
      speed up operations where many samples must be titrated, use
      an automatic buret of a type in which rubber does not come in
      contact with the solution.)

      Standard sodium thiosulfate titrants, 0.0100 N and 0.0250 N_ are"
      equivalent, respectively, to 35^-5 Vg and 886.3 yg available
      Cl/1.00 ml.

                                 3-365

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Starch indicator solution:   to 5 g starch (potato,  arrowroot,  or  soluble),
      add a little cold water and grind in a mortar to a thin  paste.
      Pour into 1 & of boiling distilled water, stir,  and let  settle
      overnight.  Use the clear supernate.  Preserve with 1.25 g  sali-
      cylic acid, h g zinc chloride,  or a combination  of h g sodium
      propionate and 2 g sodium azide/& starch solution.  Some commer-
      cial starch substitutes are satisfactory.

Standard iodine solution, 0.1 N_:  dissolve ho g KI  in  25 ml distilled
      water, add 13 g resublimed iodine, and stir until dissolved.
      Transfer to a 1-& volumetric flask and dilute to the mark.

      Standardization:  accurately measure kO to 50 ml 0.1 N_ arsenite
      solution into a flask and titrate with the 0.1 N_ iodine  solution,
      using starch solution as an indicator.  To obtain accurate
      results, it is absolutely necessary that the  solution be saturated
      with C02 at the end of the titration.  A current of C02  may be
      passed through the solution for a few minutes just before the
      end point is reached or a few drops of HC1 may be added  to
      liberate sufficient C02 to saturate the solution.

Standard iodine titrant, 0.0282 N_:  dissolve 25 g KI in a small volume
      of distilled water in a l- volumetric flask. Add 282 ml 0.1 N
      iodine solution and dilute to 1 .  Standardize  this solution
      daily with arsenite solution.  Store in amber bottles or in the
      dark.  Protect the solution from direct sunlight at all  times.
      Do not allow the solution to contact rubber.
Procedure

            Add 250 ml of a water sample to a 500-ml brown glass-
stoppered bottle or a 5QO-ml Erlenmeyer flask.  A separate blank

consisting of 250 ml of chlorine-free water should  be  prepared and

treated as a sample.
            Pipet a standardized chlorine solution  into the water

sample with rapid stirring to ensure instantaneous  mixing.   The

chlorine solution should be standardized on the day of use and of
sufficient strength so that a sample size of 15 ml  or  less can be

used to minimize any dilution effects.
            The chlorinated sample should preferably be kept in the
dark to avoid photodecomposition of the added chlorine.  Continue to

stir the solution for 15 min.
            After 15 min, withdraw a 25-ml subsample.   Acidify the

subsample to pH 3 to k with the addition of 5 ml of glacial acetic

acid and add approximately 1 g KI.  Mix the sample  and titrate with
standard sodium thiosulfate to a pale yellow color. Add 1 to  2 ml

starch solution and continue titrating to the disappearance of the

                                 3-366

-------
blue color .
            Record the volume of the standard  thiosulfate required to
titrate the sample and the blank.
            Repeat the subsampling and titration at  convenient time
intervals such as 30 min, ^5 min, and 60  min.
            Continue the test until the residual chlorine concentration
has stabilized.  If the residual chlorine concentration drops to zero,
the test should be terminated at that point  or the test should be
repeated with a higher initial chlorine concentration.
Calculations
            The chlorine demand of the sample  is the difference between
the residual chlorine concentration in the blank and the residual
chlorine concentration in the sample.  The chlorine  consumed at a
particular sampling interval can be calculated as:
                  PP _ (VB - VS)(N)(35.^ mg/meg)(V0)
where
        CC = chlorine consumed,  mg
        V               =
         B = volume 8263  to titrate  blank, ml
        V
         S = volume SaOa  to titrate sample, ml
         N = normality of standard SaOa  , meq/&
     35.^5 = equivalent weight of chlorine, mg/meq
        V
         o = volume of sample initially  chlorinated, ml
         1 = volume of subsample titrated, ml
            The chlorine demand of the sample  can be calculated as :
                             CD mg/Ji = 
where
     CD = chlorine demand, mg/
     CC = chlorine consumed at a specified time, mg
      V = volume of sample chlorinated,  Si
                                3-367

-------
                 Procedures for Sediment Samples (SID)2
Apparatus

Stirrer and magnetic stirring bar

500-ml Erlenmeyer flasks

50-ml burst
Reagents

Standard chlorine solution:   prepare by bubbling chlorine gas through
      distilled water or by diluting household bleach,  a hypochlorite
      solution, to a suitable concentration.   The concentration of the
      chlorine solution should be sufficiently strong so that the
      volume of the sample will not be increased by.more than 5 percent
      after the addition of the chlorine solution.   The useful life of
      the solution can be extended by storage in a dark or brown glass-
      stoppered bottle.  However, a chlorine  solution is unstable and
      must be standardized each time it is used.

      In order to standardize the chlorine solution, place 2 ml glacial
      acetic acid and 25 ml distilled water in a flask.  Add approxi-
      mately 1 g KI.  Pipet a convenient volume of chlorine solution
      into the flask.  Titrate with a 0.025 N. sodium thiosulfate
      solution to a pale yellow color.  Add 1 to 2 ml of starch
      indicator solution and continue the thiosulfate titration to
      the disappearance of the blue color. A blank consisting of
      2 ml of glacial acetic acid, 25 ml of distilled water, and 1 g
      KI should also be titrated to correct for any chlorine demand
      or residual chlorine in the water or reagents.
      It will be necessary to conduct a blank titration to correct the
      results for reagent impurities such as:  (a)  the free iodine
      or iodate in the potassium iodide that  liberates extra iodine or
      (b) the traces of reducing agents that  might reduce some of the
      iodine liberated.  Take a volume of distilled water corresponding
      to the sample used for titration during the standardization
      procedure, add 2 ml acetic acid, 1 g KI, and 1 ml starch solution.
      Since the blank may be either positive  or negative, it will be
      necessary to perform either (a) blank titration A or (b_) blank
      titration B, whichever applies:
      a_.  Blank titration A:  if a blue color develops, titrate with
          0.01 N_ or 0.025 N. sodium thiosulfate to the disappearance of
          the color and record results.
      b_.  Blank titration B:  if no blue color occurs,  titrate with
          0.0282 N_ iodine solution until a blue color appears.  Back-
          titrate with 0.01 N_ or 0.025 N. sodium thiosulfate and record
          the difference as titration B.
      Before calculating the chlorine consumed, subtract blank titration
      A from the sample titration, or, if necessary, add the net equi-
      valent value of blank titration B.

                                 3-368

-------
Glacial acetic acid, CH3COOH.

Potassium iodide crystals, KI.

Standard sodium thiosulfate, 0.1 N_:  dissolve 25 g Na2S203   5H20 in
      1  freshly toiled distilled water and standardize the  solution
      against potassium Mniodate or potassium dichromate after at
      least 2 weeks storage.  Use boiled distilled water and  add a
      few milliliters CHC13 to minimize bacterial decomposition of
      the thiosulfate solution.
      Standardize the 0.1 N. sodium thiosulfate titrant using  either
      (a) the Mniodate method or (b_) the dichromate method:

      a_.  Biniodate method:  dissolve 3.2^9 g primary standard quality
          anhydrous potassium Mniodate, KH(l03)2, in distilled water
          and dilute to 1 I to yield a 0.1000 N_ solution.  Store in a
          glass-stoppered bottle.
          To 80 ml distilled water, add, with constant stirring, 1 ml
          cone. H2SO^, 10 ml 0.100 N_KH(l03)2, and 1 g KI. Titrate
          immediately with sodium thiosulfate titrant until the yellow
          color of the liberated iodine is almost discharged.   Add 1 ml
          starch indicator solution and continue the titration until
          the blue color disappears.
      b_.  Dichromate method:  dissolve U.90U g primary standard quality
          anhydrous potassium dichromate, K2Cr207, in distilled water
          and dilute to 1000 ml to yield a 0.1000 N solution.   Store
          in a glass-stoppered bottle.
          Proceed as in the biniodate method, with the following
          exceptions:   substitute 10.00 ml 0.1000 N K2Cr207 for the
          KH(I03)2 and let the reaction mixture stand for 6 min in the
          dark before titrating with the Na2S203 titrant.

      The normality of the thiosulfate titrant can be calculated as:

                             Normality Na2S203 =

             (Volume oxidizing agent)(Normality oxidizing agent)
                          (Volume Na2S203 consumed)

Dilute sodium thiosulfate titrant, 0.01 N_ or 0.025 N.:  dilute 100 ml
      CO.01 N) or 250 ml (0.025 N) of 0.1 N sodium thiosulfate to 1 
      with distilled water.  To improve the stability of these titrants,
      the standard thiosulfate should be aged several weeks and the
      distilled water should be fresh.

      Standardize this solution daily in accordance with the  directions
      given above, using 0.01 N or 0.025 N KH(l03)2 or K2Cr207.  (To
      speed up operations where many samples must be titrated, use an
      automatic buret of a type in which rubber does not come in con-
      tact with the solution.)

      Standard sodium thiosulfate titrants, 0.0100 N_ and 0.0250 N_, are
      equivalent, respectively, to 35^-5 yg and 886.3 yg available
      Cl/1.00 ml.

                                  3-369

-------
Starch indicator solution:  to 5 g starch (.potato, arrowroot, or soluble).
      add a little cold water and grind in a mortar to a thin paste.
      Pour into 1  of "boiling distilled water, stir, and let settle
      overnight.  Use the clear supernate.  Preserve with 1.25 g sali-
      cylic acid, k g zinc chloride, or a combination of k g sodium
      propionate and 2 g sodium azide/Ji starch solution.  Some commer-
      cial starch substitutes are satisfactory.

Standard iodine solution, 0.1 W_:  dissolve ko g KI in 25 ml distilled
      water, add 13 g resublimed iodine, and stir until dissolved.
      Transfer to a l- volumetric flask and dilute to the mark.

      Standardization:   accurately measure ^0 to 50 ml 0.1 N_ arsenite
      solution into a flask and titrate with the 0.1 N_ iodine solution,
      using starch solution as an indicator.  To obtain accurate
      results, it is absolutely necessary that the solution be
      saturated with COa at the end of the titration.  A current of
      COz may be passed through the solution for a few minutes just
      before the end point is reached; or a few drops of HC1 may
      be added to liberate sufficient COa to saturate the solution.

Standard iodine titrant, 0.0282 N_:  dissolve 25 g KI in a small volume
      of distilled water in a l- volumetric flask.  Add 282 ml 0.1 N_
      iodine solution and dilute to 1 .  Standardize this solution
      daily with arsenite solution.  Store in amber bottles or in
      the dark.  Protect the solution from direct sunlight at all
      times.  Do not allow the solution to contact rubber.
Procedure

          Add 250 ml of chlorine-free water to a 500-ml brown glass-
stoppered bottle or a 500-ml Erlenmeyer flask.  Transfer 1 to 2 g of

well-mixed, wet sediment (SID) to the bottle.  A separate blank con-

sisting of 250 ml chlorine-free water should be prepared and treated

as a sample.
          Pipet a standardized chlorine solution into the sediment

suspension with rapid stirring to ensure instantaneous mixing.  The
chlorine solution should be standardized on the day of use and of

sufficient strength so that a sample size of 15 ml or less can be
used.  This will minimize any dilution effects.
          The chlorinated sample should preferably be kept in the

dark to avoid photodecomposition of the added chlorine.  Continue
to stir the sediment suspension for 15 min.
          After 15 min, withdraw a 25-ml subsample.  Acidify the

subsample to pH 3 to 4 with the addition of 5 nil glacial acetic acid
and add approximately 1 g KI.  Mix the sample and titrate with

standard thiosulfate to a pale yellow color.  Add 1 ml starch solution

                                 3-3TO

-------
and continue titrating to the disappearance of the blue color.   Starch
may have to be added immediately to the sediment suspension  if  the yel-
low iodine color is not visible due to turbidity.
            Record the volume of thiosulfate required  to titrate the
sample and the blank.
            Repeat the subsampling and titrations  at convenient time
intervals such as 30 min, U5 min, and 60 min.
            Continue the test until the residual chlorine concentration
has stabilized.  If the residual chlorine concentration drops to zero,
the test should be terminated at that time or the  test should be
repeated using a higher initial chlorine concentration.
Calculations
            The amount of chlorine consumed at a specified sampling
interval can be calculated as:
                  rr   (VB - VS)(N)(35.U5 mg/meq)(Vo)
                  CC "              (V!)
where
        CC = chlorine consumed, mg
        V
         B = volume thiosulfate to titrate blank,  ml
        V
         S = volume thiosulfate to titrate sample, ml
         N = normality of thiosulfate, meq/
     35.^5 = equivalent weight of chlorine, mg/meq
        ^ = volume of distilled water in sample,  ml
        Vi = volume of subsample titrated, ml
            The chlorine demand of the sediment is calculated as:
                  CD mg/kg (wet weight) = (CC)(l000)
                                               o
                  CD mg/kg (dry weight) =
where
      CD = chlorine demand, mg/kg
      CC = chlorine consumed at a specified time,  mg
       g = wet weight of sediment sample chlorinated,  g
     % S = percent solids in sediment sample (expressed  as a decimal
           fraction)
                                 3-371

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                              References
1.  American Public Health Association.   Standard Methods for the
    Examination of Water and Wastewater  Including Bottom Sediments
    and Sludges. iHh Edition.   APHA;  New York,  New York.  1193 p.
    (1975).
2.  Great Lakes Region Committee on Analytical Methods.   "Chemistry
    Laboratory Manual for Bottom Sediments."  U. S. Department of
    the Interior, Great Lakes Basin; Chicago,  Illinois.   96 p. (1968)
                                 3-372

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                       BIOCHEMICAL OXYGEN DEMAND


            The biochemical oxygen demand (BOD) test is an emperical

bioassay type procedure that measures the dissolved oxygen (DO) consumed

by microbial organisms while assimilating and oxidizing the organic

matter present.  The procedure consists of measuring the change in

oxygen concentration in a sample during a 5-day period at 20C and in

the dark.  Although it is generally realized that 20 days or longer

may be required to completely stabilize the organic material in an

environmental sample, the 5-day period has been accepted as standard

because of practical considerations and the fact that a large percentage

of the ultimate demand is satisfied in the first 5 days.

            Three methods have been used to measure the BOD of a sample.

The selection between these methods depends on the amount of oxygen

consumed during organic stabilization as indicated below:
            a_.  Direct method:  used with samples whose 5-day BOD is
                determined by measuring the dissolved oxygen content
                of the water before and after a standard incubation
                period of 5 days at 20C.  Samples are aerated at the
                start of the test to raise dissolved oxygen concen-
                trations to near saturation.  The change in oxygen
                is based on the microbial population of the sample
                and additional test organisms are not added to the
                sample.
            b_.  Unseeded dilution method:  used with waters having BOD
                values greater than 7 mg/.  Sample aliquots are di-
                luted using water saturated with oxygen (dilution
                water).  The dissolved oxygen concentration is deter-
                mined immediately after dilution and after 5 days
                incubation at 20C.
            c_.  Seeded dilution method:  used with samples having low
                BOD values.  When the microbial population of the
                sample is low or potentially toxic conditions exist
                in the sample, a mixed group of organisms, commonly
                called a seed, is added to the sample.  Oxygen con-
                centrations are then determined on the initial sample
                and after incubating for 5 days at 20C.


                      Sample Handling and Storage


            The BOD test can be performed with either water or sediment

                                 3-373

-------
samples.  In either case, the sample should be protected from contact

with atmospheric oxygen, and the analysis should be initiated within

k to 6 hr of collection.  This requires that wet sediments should be

used and that both sediment and water samples to be analyzed for BOD
be shipped and stored in airtight containers to minimize oxidation

(Figure 3-^5).
            There is no recommended preservative at this time other

than refrigeration at ljC and a short holding time.

                                                         3
            Procedures for Water Samples (Wl, W2, S1A)1 '2


Apparatus
Incubation bottles, 300-ml bottles with ground -glass stoppers
Incubator, thermostatically controlled at 20 C +_ 1C:   all light
      should be excluded to prevent photosynthetic production of
      dissolved oxygen by algae in the sample
Graduated cylinders, 1 H or 2 &

Reagents
Distilled water:   water used for solutions and for preparation of the
      dilution water must be of highest quality, distilled from a
      block tin or all-glass still, contain less than  0.01 mg/
      copper, and be free of chlorine, chloramines, caustic alkalinity,
      organic material, or acids.
Phosphate buffer solution:  dissolve 8.5 g potassium dihydrogen
      phosphate, KHzPOi*; 21.75 g dipotassium hydrogen  phosphate,
              33.it g disodium hydrogen phosphate heptahydrate,
               7HaO; and 1.7 g ammonium chloride, NHifCl, in about
      500 ml distilled water and dilute to 1 H.   The pH of this
      buffer should be 7.2 without further adjustment.  If dilution
      water is to be stored in the incubator, the phosphate buffer
      should be added just prior to using the dilution water.

Magnesium sulfate solution:  dissolve 22.5 g magnesium sulfate,
      MgSOn  7H20, in distilled water and dilute to 1 .
Calcium chloride solution:  dissolve 27  5 g anhydrous  calcium chloride
             in distilled water and dilute to 1 &.
Ferric chloride solution:   dissolve 0.25 g ferric chloride, FeCla *
      6H20, in distilled water and dilute to 1 .
Acid and alkali solutions, 1 N_:  for neutralization of waste samples
      which are either caustic or acidic.

* References for this section are found on page 3-38U.

                                 3-37^

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      Figure  3-^5.      Handling  and storage  of  samples  for  "biological oxygen  demand analysis
" TR(ENT




1
ELUTRIATE

STORE WET



+
DIGEST
        I
                                                                                           1
SAMPLE
DESIGNATION
W1
W2
W3
S1A
S1B
SIC
SID
S2
S3
PURPOSE
CONTAINER
SAMPLE TREATMENT
PRESERVATIVE
STORAGE TIME
DIGESTION SOLUTION
SAMPLE VOLUME OR WEIGHT
                         Total Water
                            Cone.
                            G,P
                            None
                           6  hr
Soluble
 Water
 Cone.

 G,P
                                        ltC
                                       6 hr
                          0.3-U       0.3-U
 Used in
Elutriate
   G,P
                                        Filter      None
                                                   None
                                                   <1 w
Mobile
 Cone.
  G.P
                                                                 None
  Total
Sediment
  Cone.

   G,P
                                                                                                    None
                          
-------
Sodium sulfite solution, 0.025 N_:  dissolve 1.575 g anhydrous sodium
      sulfite, Na2S03, in 1 Si distilled water.  This solution is not
      stable and should be prepared daily.

Seeding material:  satisfactory seed may sometimes be obtained by using
      the supernatant liquor from domestic sewage which has been stored
      at 20 C for 2h to 36 hr.  Refer to ASTM3 for a more detailed
      explanation of seeding material.

Dilution water:  the distilled water used should have been stored in
      cotton-plugged bottles for a sufficient length of time to
      become saturated with DO.  The water should be aerated by
      shaking a partially filled bottle or using a supply of clean
      compressed air.  Situations may be encountered where it is
      desired to use stabilized river water to check stream perfor-
      mance with laboratory procedure.  The distilled water used
      should be as near as possible to 20C and of the highest purity.
      Place the desired volume of distilled water in a suitable
      bottle and add 1 ml each of phosphate buffer, magnesium
      sulfate, calcium chloride, and ferric chloride solutions
      'for each liter of water.

      Seeding:  if necessary, the dilution water is seeded using the
      seed found to be the most satisfactory for the particular
      material under study.  Only past experience can determine the
      actual amount of seed to be added per liter.  Seeded dilution
      water should be used the same day it is prepared.

Sample pretreatment

            Samples containing acidity or caustic alkalinity are
neutralized to approximately pH 7 with H2SOit(l H_) or NaOH(l N_) using
a pH meter.

            Chlorine residuals, if present in a sample, may dissipate
after 1 to 2 hr.   If chlorine residuals do not dissipate on standing,

the neutralized sample must be treated with 0.025 K_ sodium sulfite

solution.  The appropriate quantity of sodium sulfite solution is
determined on a 100- to 1000-ml portion of the sample by adding 10 ml

1 + 1 acetic acid or 1 + 50 HjSOit, followed by 10 ml potassium iodide

solution (10 g in 100 ml) and titrating with 0.025 N sodium sulfite

solution to the starch-iodide endpoint.  Add to a volume of sample
the quantity of sodium sulfite solution determined by the above test;
mix; and after 10 to 20 min, test sample aliquots for residual

chlorine to check the treatment.
            Samples supersaturated with oxygen must be reduced to

saturation (9-17 mg/Jl at 20C) by aerating with compressed air or by

vigorous shaking of the sample container.

                                 3-376

-------
            The sample pretreatment steps outlined only apply to water
samples.  It is expected that these sample preparation techniques will
be required seldom, if ever, with sediment samples.
Procedure
            Direct method for water.  Fill two BOD bottles with the
Wl, W2, or S1A sample.  Be sure that no air bubbles are entrapped and
that the bottles are filled to overflowing when the stoppers are
inserted.
            Determine the dissolved oxygen concentration in one of the
bottles.  Record the value as initial DO.
            Allow the other bottle to incubate for 5 days in the dark
at 20C and then determine the dissolved oxygen content.  Record the
value as final DO.
            For the direct method, calculate the BOD as follows:
                   BOD rag/a = Initial DO - Final DO
            Unseeded dilution method for water.  Conduct any sample
pretreatments that are necessary.
            Fill a 1000-  to 2000-ml capacity graduated  cylinder  approxi-
mately half full with dilution water.  Add the volume of carefully
mixed sample to produce the desired final dilution and dilute to the
mark with dilution water.  Mix well with a plunger-type mixing rod,
taking precautions to avoid any entrainment of air.  It is suggested
that a minimum of three dilutions be prepared of each sample.
NOTE:  Dilutions may also be prepared by directly pipetting known
       volumes of the sample into BOD bottles, providing the volume
       of the BOD bottle is known.  However, this method should not
       be used if dilutions of 100:1 or more are required.
            Determine the dissolved oxygen in one of the BOD bottles
prepared above.  Record as initial DO.
            Incubate one of the remaining bottles for 15 min in the
dark at 20C and determine the oxygen content after the incubation for
calculation of the immediate oxygen demand (IDOD).  Record result as
DO after 15 min.
            Incubate the remaining bottle for 5 days and then deter-
mine the oxygen content.   Record as final DO.
                                3-377

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            When the. unseeded dilution method is used,  the BOD of the
sample is calculated as follows:
                         BOD mg/ = (I-F)0>)
where
     I = dissolved oxygen concentration after 15 min, mg/
     F = dissolved oxygen concentration after 5 days, mg/Jl
     b = volume of BOD bottle, ml
     v = volume of sample BOD bottle, ml
            Seeded dilution method.  The procedure used in the unseeded
dilution method is followed; but  an additional step is  necessary and
that is to correct for the effect of the seed depletion of DO.  Deter-
mine the oxygen depletion of the  seed by setting up a separate series
of seed dilutions (controls) and  selecting those resulting in a kO per-
cent to 70 percent depletion in 5 days.  One of these depletions is
then used to calculate the correction due to the small  amount of seed
in the dilution water.
            Fill a 1000-  to 2000-ml capacity graduated  cylinder  approxi-
mately half full with seeded dilution water.  Add the volume of care-
fully mixed sample to produce the desired final dilution and dilute
to the mark with, dilution water.   Mix well with a plunger-type mixing
rod, taking precautions to avoid  any entrainment of air.   It is sug-
gested that a minimum of three dilutions be prepared of each sample.
NOTE:  Preparation of diluted samples may also be accomplished by
       direct measurement of suitable amounts of sample into BOD
       bottles using a large-tipped volumetric pipette  and then
       filling the bottles with dilution water.  The volume of each
       bottle will have to be measured in order to calculate dilution
       factors needed to determine seed corrections. Dilutions
       greater than 100:1 must be performed in graduated cylinders.
            Siphon, with continued mixing, the diluted  sample to
completely fill three bottles.  One bottle is for the determination
of initial dissolved oxygen concentration.  The second  bottle is
incubated for 15 min and used to  determine immediate dissolved oxygen
demand (IDOD).  The third sample  is incubated for 5 days at 20C and
analyzed to determine oxygen consumption.
                                3-378

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Calculations
            The seed correction factor is calculated as:
                                     y
where
     C = seed correction factor, mg/
     B = BOD of seed control, mg/
     x = percent seed in sample
     y = percent seed in control
            The BOD of the sample is calculated as:
                       BOD
where
     I = dissolved oxygen concentration after 15 min,  mg/&
     F = final dissolved oxygen concentration after  5  days,  mg/
     C = seed correction factor
     v = volume of sample in BOD bottle, ml
     b = volume of BOD bottle, ml
            The immediate dissolved oxygen demand (IDOD)  is  calculated
as:
where
     0 = dissolved oxygen concentration at time zero, mg/
     I = dissolved oxygen concentration after 15 min,. mg/
     v = volume of sample in BOD bottle, ml
     b = volume of BOD bottle, ml
                                3-379

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                Procedures for Sediment Samples (SID)1*


Apparatus

Incubation bottles, 300-ml capacity, vith ground glass stoppers

Incubator, thermostatically controlled at 20  +_ 1C:   all light
      should be excluded to prevent the photosynthetic  production
      of dissolved oxygen by algae in the sample

Graduated cylinders, 1  or 2 

Reagents

Distilled water:  free of copper, chlorine, chloramines, caustic
      alkalinity, acids, and organic material.

Phosphate buffer solution:  dissolve 8.5 g potassium dihydrogen
      phosphate, K^PO^ 21.75 g dipotassium hydrogen phosphate,
      K2HP(\; 33.k g disodium hydrogen phosphate heptahydrate,
      Na2HP04   7H20; and 1.7 g ammonium chloride, NHi,Cl, in distilled
      vater and dilute to 1 .  The pH of this buffer should be 7.2
      without further adjustment.  If dilution water is to be stored
      in the incubator, the phosphate buffer should be  added just
      prior to using the dilution water.

Magnesium sulfate solution:  dissolve 22.5 g magnesium sulfate,
      MgS(\  7H20, in distilled water and dilute to 1  .

Calcium chloride solution:  dissolve 27-5 g anhydrous calcium
      chloride,  CaCl2, in distilled water and dilute to 1 .
Ferric chloride solution:  dissolve 0.25 g ferric chloride, FeCls 
      6H20, in distilled water and dilute to 1 .

Sodium sulfite solution, 0.025 N.:  dissolve 1.575 g anhydrous sodium
      sulfite, Na2S03, in 1  distilled water.  This solution is not
      stable and should be prepared daily.
Seeding material:  satisfactory seed may sometimes be obtained by
      using the supernatant liquor from domestic sewage which has
      been stored at 20C for 2U to 36 hr.  Refer to ASTM3 for a
      more detailed explanation of seeding material.  .
Dilution water:   store distilled water in cotton-plugged bottles for
      a sufficient length of time to become saturated with DO.  The
      water should be aerated by shaking a partially filled bottle
      or using a supply of clean compressed air.  Situations may be
      encountered where it is desired to use stabilized river water
      to check stream performance with laboratory procedure.  The
      distilled water used should be as near as possible to 20C and
      of high purity.  Place the desired volume of distilled water
      in a suitable bottle and add 1 ml each of phosphate buffer,
      magnesium sulfate, calcium chloride, and ferric chloride for
      each liter of water.
      Seeding:  use the seed that has been found by practical experi-
      ence to be the most satisfactory for the particular material
                                3-380

-------
      under study.  Only past experience can determine the actual amount
      of seed to be added per liter but the amount should give an oxygen
      depletion of approximately 2 mg/.  The amount of seed required
      may vary with the source of the seed and will have to be established
      through experience.  If the sample contains organic compounds not
      amenable to oxidation by domestic sewage seed, it may be necessary
      to use seed prepared from soil, an acclimated seed developed in the
      laboratory, or sediments collected below a particular waste dis-
      charge (preferably 2 to 5 miles below the point of discharge).
      Seeded dilution water should be used the same day it is prepared.
Procedure
            Weigh an appropriate size SID sediment sample directly into
the BOD bottle (suggested weight of 0.5 to 5.0 g).  Each sample should
be prepared in replicate.
            Fill each BOD bottle with dilution water and place the
samples in the incubator.  Ensure that air bubbles are not trapped in
the BOD bottles.  Prepare a blank consisting of dilution water in a
separate BOD bottle.  Make sure that there is a water seal in the neck
of each sample bottle and blank when placed in the incubator.  Replenish
the water seals on all bottles each morning.
            Determine the initial dissolved oxygen concentration of the
sample using the azide modification of the iodometric method or a
dissolved oxygen probe.  This can best be accomplished by directly
measuring the dissolved oxygen concentration in the dilution water.
This method is recommended because sediment may cause a rapid con-
sumption of oxygen, making it difficult to obtain a stable initial
dissolved oxygen reading.  (if a probe is used for oxygen measurement,
the same sample can be used for immediate dissolved oxygen demand and
biochemical oxygen demand.)
            Incubate a blank (dilution water) and the sediment sus-
pensions for 5 days at 20C.  Determine residual dissolved oxygen
concentrations in the incubated samples using the analytical method
of choice.   The most reliable BOD determinations will occur in those
samples with a residual DO of at least 2 mg/ and a DO depletion of
at least 2 mg/.
            It may be desirable to incubate the dilution water as a
check on its quality.  In order to do this, fill two BOD bottles with
unseeded dilution water.  Stopper one bottle, fill the water seal,
                                 3-381

-------
and place in the incubator for 5 days.   Analyze the second sample to
determine initial dissolved oxygen concentration.   Following the 5-day
period, determine dissolved oxygen in the incubated sample.   The oxygen
depletion should not be more than 0.2 mg/ and preferably not more than
0.1 mg/.  If these values are exceeded, the quality of the  dilution
water or the treatment of samples (filling of water seals, etc.) should
be considered suspect.
            It is also recommended that the analyst routinely run pure
organic compounds for which the BOD is known or determinable.  This is
necessary because the quality of the dilution water, the effectiveness
of the seed, the technique of the analyst, and the presence of toxic sub-
stances can all influence BOD results.   The use of known standards will
indicate whether any of the identified factors are out of control.
            Prepare a stock BOD standard solution  by dissolving 0.150 g
reagent grade glucose and 0.150 g reag'ent grade glutamic acid in 1  of
distilled water.  The solids should be dried for 1 hr at 103C prior
to weighing.
            Prepare a working BOD standard solution by diluting 20 ml
of the stock solution to 1  with seeded dilution  water.  Fill three
BOD bottles and incubate at 20C for 5  days.  The  resultant  BOD of
these samples should be 2l8 mg/ +_ 11 mg/.  Any appreciable deviation
from these expected results raises questions on the quality  of the
dilution water, the viability or suitability of the seed material,
or the analytical technique.
Interferences
            Many synthetic organic components in industrial  waste
waters and sediments are not biodegradable without the seeding pro-
cedure because of either a toxic effect or a deficiency or absence of
appropriate microorganisms.
            Chlorine residuals must be removed prior to the  test
because residual chlorine may be toxic  to the microbial population
or may oxidize organic material.
            Because waters and sediments that contain sulfide, sulfite,
or ferrous ions create an immediate demand on the  dissolved  oxygen,
it is necessary to distinguish this immediate dissolved oxygen demand

                                  3-382

-------
ClDOD) from the true BOD.  The depletion of DO in a standard water dilu-
tion of the sample in 15 min has been arbitrarily selected as the IDOD.
Calculations
            Immediate dissolved oxygen demand (IDOD) is calculated as
follows:
                 IDOD mg/kg (wet weight) =

                 IDOD mg/kg (dry weight) =
where
       0 = dissolved oxygen concentration at time zero, mg/&
       I = dissolved oxygen concentration after 15 min, mg/
       b = volume of BOD bottle, ml
       g = wet weight of sediment sample used, g
     % S = percent solids in sediment sample (expressed as a decimal
           fraction)
            The sediment BOD is calculated as follows:
                  BOD mg/kg (wet weight) =

                  BOD mg/kg (dry weight) =
where
       0 = dissolved oxygen concentration at time zero, mg/Jl
       F = dissolved oxygen concentration after 5 days, mg/&
       b = volume of BOD bottle, ml
       g = wet weight of sediment sample used, g
     % S = percent solids in sediment sample (.expressed as a decimal
           fraction)
                                 3-383

-------
                              References
1.  American Public Health Association.   Standard Methods for the Exami-
    nation of Water and Wastewater Including Bottom Sediments and Sludges,
    lUth. Edition.  APHA; New York, New York.  1193 p. (1975).
2.  Environment Canada.  "Analytical Methods Manual."  Inland Waters
    Directorate, Water Quality Branch; Ottawa, Ontario, Canada (l9T^)-
3.  American Society for Testing and Materials.  Book of ASTM Standards.
    Part 31. Water.  American Society for Testing and Materials;
    Philadelphia, Pennsylvania (1976).
i|.  Great Lakes Region Committee on Analytical Methods.  "Chemistry
    Laboratory Manual for Bottom Sediments."  U. S. Department of the
    Interior, Great Lakes Region; Chicago, Illinois.  96 p.  (1968).
                                 3-38U

-------
                        CHEMICAL OXYGEN DEMAND

            The chemical oxygen demand (COD)  test was devised as  an alter-
nate to the biochemical oxygen demand test for estimating organic matter.
The procedure consists of digesting a sample  with a strong oxidizing
agent at elevated temperatures and reduced pH.  The amount of oxidizing
agent consumed during the test is expressed as an equivalent amount of
oxygen.  Since most organic compounds will be oxidized under conditions
of the test, results are considered a measure of the amount of oxygen
required to stabilize organic matter present  in the sample.
            The COD procedure can be used to  characterize a sample.
However, the user is cautioned that the method is not specific for
organic matter.  A number of inorganic substances that may be present
                    +2    +2   -2    -
in water samples, Fe  , Mn  , S  , NO 2, can increase the  consumption
of oxidizing agent during the test.  As a consequence, a  lack of  cor-
relation between COD results and other tests  that measure organic
carbon (BOD, TOG) has been reported.  This problem will be amplified
in sediment samples due to the reduced nature of most sediments and
the higher concentration of reduced inorganic species such as Fe  z,
   2        2
Mn  , and S  .   It is recommended that COD results not be equated with
organic matter in sediments.

                      Sample Handling and Storage

            Information for the handling of COD samples is summarized
in Figure 3-^6.  Water samples may be stored  in either glass or plastic
containers and preserved for up to 7 days with sulfuric acid.  Sediment
samples may also be stored in either glass or plastic containers.  How-
ever, since there are no chemical preservative agents and sediment COD
can be affected by air oxidation, it is suggested that only field moist
sediment samples (SID) be used for COD analysis.
                                 3-385

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       Figure  3-^6.     Handling  and storage of  samples  for chemical oxygen  demand  analysis

^
AC 1 D 1 FY
1
STORE
i
DIGEST
I
ANALYZE
(W1)


CORE SAMPLE
4
>
WATER SAMPLE DREDGE SAMPLE CORE SECTION 1
f 1,4
* * 1
FILTER HOTREATHEHT SJORE WET |
1 	
ACIDIFY
, 1
STORE
1 '
+ ^
fr ELUTRIATE DIGEST

1 1 1
ANALYZE ANALYZE ANALYZE
(W2) (S1A) (SID)

OO
CT\ SAMPLE DESIGNATION
PURPOSE
CONTAINER
SAMPLE TREATMENT
PRESERVATIVE
W1 W2 W3 S1A SIB SIC SID S2 S3
Total Water Soluble Used In Mobile Total
Cone. Water Elutriate Cone. Sediment
Cone. Cone.
G,P G,P G,P G,P G,P
None Filter None None None
H2SOi H2SOi, (Minimize Air Contact. Keep Field Moist,)
pH<2 pH<2
STORAGE TIME
                        7 d
                                  7 d
DIGESTION SOLUTION
SAMPLE VOLUME OR WEIGHT
                       20 ml
                                 20 ml
                                                      20 ml
                                                                                    2 9

-------
             Procedures for Water Samples (Wl, W2, S1A)1 '2
Method 1:  Low Level, 5 to 50 mg/
Apparatus
Reflux apparatus:  consisting of 250- or 500-ml Erlenmeyer flasks
      with ground-glass 2U/1+0 neck** and 300-mm jacket Liebig, West, or
      equivalent condenserst with 2^AO ground-glass joint

Hot plate:  having sufficient power to produce I.k W/cm2 (9 W/in.2) of
      heating surface, or equivalent, to ensure adequate refluxing of
      the sample
Reagents
Standard potassium dichromate solution, 0.250 N_:  dissolve 12.259 g
      potassium dichromate, K2Cr207, primary standard grade, previously
      dried at 103C for 2 hr, in distilled water and dilute to 1 H.
      To eliminate the interference of nitrites, sulphamic acid, in the
      amount of 10 mg for every 1 mg of nitrite N in the refluxing
      flask, may be added to the dichromate solution.  Thus, 0.12 g/
      sulphamic acid added to the dichromate solution will eliminate
      the interference of nitrites up to 6 mg/& in the sample.
Dilute potassium dichromate solution, 0.025 N.:  dilute 100 ml of
      standard potassium dichromate solution, 0.250 N, to 1  with
      distilled water.
Sulfuric acid reagent:  cone. H2SOi, containing 22 g silver nitrate,
      Ag2SOit, per 9-lb bottle (l or 2 days required for dissolution).

Standard ferrous ammonium sulfate titrant, 0.25 N_:  dissolve 98 g
      ferrous ammonium sulfate, Fe(NH4 )2(SOil )a * 6H20, in distilled
      water.  Add 20 ml of cone. H2S04 (CAUTIONI) and allow to cool.
      Titrate with the ferrous ammonium sulfate titrant, using 2 or 3
      drops of ferroin indicator.
                     Normalitv = fo1 29^07X0.25)
                     Normality   Lml Fe(NHi|)2(SOlt)2]
Standard ferrous ammonium sulfate, 0.025 N_:  dilute 100 ml ferrous
      ammonium sulfate, 0.25 N_,to 1  with distilled water.  This
      solution must be standardized daily against the dilute potassium
      dichromate solution, 0.025 N., following the same procedure as
      the standardization of the ferrous ammonium sulfate titrant,
      0.25 1-
Ferroin indicator solution:  dissolve 1.U85 g 1,10-phenanthroline
      monohydrate and 0.695 g ferrous sulfate, FeSOi*  TH20, in water
      and dilute to 100 ml.  Alternatively, a commerically prepared
      indicator can be purchased.

 * References for this section are found on page 3-395.
** Corning 5000 or equivalent.
 t Corning 2360, 915^8, or equivalent.

                                  3-38T

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Silver sulfate, AgaSCK, reagent powder.
Mercuric sulfate, HgSOi*, analytical-grade crystals.
Procedure
            Place several boiling stones or glass beads in the reflux
flask.  Add 20.0 ml of sample or an aliquot diluted  to 20.0 ml.  Add
O.U g HgSOit to the reflux flask and mix.  (The O.U g HgSOi is sufficient
to complex ^0 mg chloride ion or up to 2000 mg/& in  a 50-ml sample.
If the sample chloride concentration exceeds this value, additional
HgSOit must be added to maintain a HgSOitiCl ratio of  10:1.)
            Cool the reflux flask in an ice bath and slowly add 10 ml
0.025 N. K2Cr207.  The sample should be continuously  mixed or swirled
during this step.
            Add 30 ml of sulfuric acid-silver sulfate reagent to the
cooled solution.  This addition should be performed  slowly and with
constant sample swirling for two reasons:  the combination of ice
bath temperatures and slow addition of the sulfuric  reagent is intended
to minimize the loss of volatile organic compounds;  and the thorough
mixing of acidified samples is intended to prevent local heating that
can result in superheating and the sample being blown out the con-
denser during reflux.
            Attach sample flask to the condenser; start the cooling
water and reflux for 2 hr.
            Allow the flask to cool and wash down the condenser with
25 to 30 ml distilled water.  If the reflux flask has a flat bottom,
the final titration may be run in the same flask. If a round-bottomed
flask has been used, quantitatively transfer the sample solution to  a
250-ml Erlenmeyer flask, washing out the reflux flask 3 or ^ times with
distilled water.  After the sample has reached room  temperature, add
3 drops of ferroin indicator.  The quantity of ferroin indicator used
on all samples should be consistent.  Titrate the excess dichromate
with 0.025 N, ferrous ammonium sulfate.  The endpoint of the titration
will be indicated by a sharp color change from blue-green to reddish-
brown .
            A blank consisting of 20 ml distilled water is to be
processed as a sample to check for reagent contamination.
                                3-388

-------
Interferences

            Traces of organic material from the glassware or the atmos-
phere may cause a positive error in the COD test.
            Care should be exercised to avoid inclusion of organic
materials in distilled water used for reagent preparation or sample
dilution.
            Glassware used in the test should be conditioned by running
blank procedures to eliminate traces of organic material.
            Volatile materials may be lost as the sample temperature
rises during the addition of sulfuric acid and reagent.  This loss
can be minimized by cooling the sample flask during this step.
            Chlorides are quantitatively oxidized by dichromate and
represent a positive interference in the COD procedure.  Mercuric
sulfate is added to complex chlorides.  The mercuric sulf ate: chloride
ratio should be at least 10:1 to minimize this interference.
Calculations
            The COD of the sample is calculated as follows:
                                 (A-B)(N)(8000)
                      COD
                                         S
where
        A = volume FeCNHit )2 (SO.* )2 used for blank titration, ml
        B = volume Fe(NHt|)2 (SOt^ )2 used for sample titration, ml
        N = normality of Fe(NHH )2(SOil )2 used, eq/&
     8000 = equivalent weight of oxygen, mg/eq
        S = volume of sample, ml

Method 2:  High Level, 50 to 800 mg/&2
            The high-level COD procedure is very similar to the low-
level COD procedure except for the strength of the dichromate solution,
the strength of the titrant, and the optional use of a chloride-
correction procedure.  When the chloride concentration in the sample
exceeds 1000 mg/, the minimum reportable COD value will be 250 mg/&
because of the large chloride-correction factor.
                                 3-389

-------
Apparatus
Reflux apparatus:  consisting of 250- or 500-ml Erlenmeyer flasks with
      ground-glass 2k/kO neck* and 300-mm jacket Liebig, West, or
      equivalent condensers** with 2U/UO ground-glass joint
Hot plate:  having sufficient power to produce l.U W/cm2 (9 W/in.2) of
      heating surface, or equivalent, to ensure adequate refluxing of
      the sample
Reagents

Standard potassium dichromate solution, 0.250 N_:  dissolve 12.259 g
      potassium dichromate,  KaCraO?, primary standard grade, previously
      dried at 103C for 2 hr, in distilled water and dilute to 1 H.
      The addition of 0.12 g/& sulphamic acid will eliminate inter-
      ference due to nitrites in the sample at concentrations up to
      6 mg/Jl.
Dilute potassium dichromate solution, 0.025 N.:  dilute 100 ml of standard
      potassium dichromate solution, 0.250 N_, to 1 & with distilled
      water.
Sulfuric acid reagents:  cone . HgSO^ containing 22 g silver sulfate,
      Ag2S0lt, per 9-lb bottle.  Allow 1 or 2 days for dissolution.
Standard ferrous ammonium sulfate titrant, 0.25 N;.  dissolve 98 g
      ferrous ammonium sulfate, Fe(NHit ) 2(80^)2  6HaO, in distilled
      water.  Add 20 ml cone. H2SOi, (CAUTION.1), cool, and dilute to
      1 .  This solution must be standardized against Kj-CraO? daily.
      Standardization of ferrous ammonium sulfate:  dilute 10 ml
      standard potassium dichromate solution to approximately 100 ml.
      Add 30 ml cone. HaSO.*  (CAUTION!) and allow to cool.  Titrate
      with the ferrous ammonium sulfate titrant, using 2 or 3 drops
      of ferroin indicator.

                    Wn,-H+v - (ml K2Cr 2.0.7) (0.25)
                    Normality - [ml Fe(HHjz(SO,)2]

Dilute ferrous ammonium sulfate, 0.025 N_:  dilute 100 ml standard
      ferrous ammonium sulfate, 0.25 N, to 1 & with distilled water.
      This solution must be  standardized daily against the dilute
      potassium dichromate solution, 0.025 N., following the same
      procedure as the standardization of the ferrous ammonium
      sulfate titrant, 0.25  N.
Ferroin indicator solution:  dissolve 1.U85 g 1,10-phenanthroline
      monohydrate and 0.695  g ferrous sulfate, FeSOi*   TH20, in water
      and dilute to 100 ml.  Alternatively, a commerically prepared
      indicator can be purchased.
Silver sulfate, AgaSOi*, reagent powder.
Mercuric sulfate, HgSOn, analytical-grade crystals.

Procedure
            Place several boiling stones or glass beads in a reflux
 * Corning 5000 or equivalent.
** Corning 2360, 915^8, or equivalent,
                                  3-390

-------
flask.  Add 20.0 ml of a sample or an aliquot diluted to 20.0 ml.   Add
HgSOi* in the ratio of 10 mg HgSOi to 1 mg chloride,  based on the mg
chloride in the sample aliquot.  (An addition of 1.0 g HgSOi will  be
sufficient to complex 100 mg chloride in the sample  aliquot.)  Swirl
to dissolve the mercuric sulfate.
            Cool the sample in an ice bath and slowly add 10 ml 0.25 N.
KaC^Oy.  To the well-mixed solution, slowly add 30  ml sulfuric acid-
silver sulfate reagent.  If a high concentration of  volatile organic
compounds is known or suspected to be present, the sulfuric  acid-
silver sulfate solution can be added through the condenser of an
Allihn condenser to reduce volatilization losses.
            Thoroughly mix the acidified sample to prevent local heating
and possible sample loss (superheated sample may be  blown out of the
condenser).  Attach the flask to a condenser and reflux for  2 hr.
            Allow the sample to cool and wash the condenser  with
25 to 30 ml distilled water.
            When the sample has reached room temperature, add 3 drops
of ferroin indicator.  Titrate the excess dichromate with 0.25 N.
ferrous ammonium sulfate solution until a sharp color change occurs
(blue-green to reddish-brown)
            A blank consisting of 20 ml distilled water must be carried
through the analytical procedure to correct for reagent contamination.
            For COD values greater than 800 mg/, a  smaller  sample
aliquot should be used.  Howeverj the volume of the  aliquot  should be
diluted to 20 ml using a distilled water-sodium chloride solution  with
a chloride concentration equal to the sample.
            Chloride correction:   When the sample chloride  concentra-
tion exceeds 1000 mg/Jl, prepare a standard curve of  COD vs.  mg/&
chloride.  This is accomplished by preparing a series of sodium
chloride solutions whose chloride concentrations bracket the chloride
concentration of the sample(s).  These solutions are processed as  COD
samples and the resultant COD's are plotted vs. chloride concentration.
Do not extrapolate beyond the upper or lower limits  of the chloride
curve.
                                  3-391

-------
Calculations
            The COD of the sample is calculated as follows:
                              - B)N x 8000] - ?0 D
              COD mg/ =
                                      o
where
        A = volume 0.25 N FefNIUMSOit h for blank titration, ml
        B = volume 0.25 N Fe(NH4 )2(SOif }z for sample titration, ml
        N = normality of Fe^OaCSOOz used for titration, eq/
     8000 = equivalent weight of oxygen, mg/eq
        S = volume of sample used in test, ml
        D = chloride correction from COD-chloride curve
      1.2 = correction factor to compensate for the different oxidation
            of chloride in organic-containing (sample) and nonorganic
            containing (NaCl-di stilled water) systems.
                                  3-392

-------
                Procedures for Sediment Samples (SID)*4
Apparatus
Reflux apparatus:  consisting of 250- or 500-ml Erlenmeyer flasks with
      ground-glass 2h/kO neck* and 300-mm jacket Liebig, West, or
      equivalent condensers** with 24/UO ground-glass joint

Hot plate:  having sufficient power to produce l.U W/cm2 (9 W/in.2) of
      heating surface, or equivalent, to ensure adequate refluxing of
      the sample

Reagents

Standard potassium dichromate solution, 0.250 N_:  dissolve 12.259 g
      potassium dichromate, KaCr^Oy, primary standard grade, previously
      dried at 103C for 2 hr, in distilled water and dilute to 1 .
      The addition of 0.12 g/Jl sulphamic acid will eliminate inter-
      ference due to nitrites in the sample at concentrations up to
      6 mg/JL
Sulfuric acid reagent:  cone. HzSOi* containing 22 g silver sulfate,
      AgaSOit, per 9-lb bottle.  Allow 1 or 2 days for dissolution.

Standard ferrous ammonium sulfate titrant, 0.250 IT:  dissolve 98 g
      ferrous ammonium sulfate, Fe(NHit )z (SOn )a   6HaO, in distilled
      water.  Add 20 ml cone. HaSOit (CAUTION!), cool, and dilute
      to 1 .  This solution must be standardized against KzC^Oy daily.
      Standardization of ferrous ammonium sulfate:  dilute 10 ml
      standard potassium dichromate solution to approximately 100 ml.
      Add 30 ml cone. E2SQn (CAUTION!) and allow to cool.  Titrate
      with ferrous ammonium titrant, using 2 or 3 drops of ferroin
      indicator.

                    w __ , , .+  _ (ml K2Cr207)(0.25)
                    Normality - tml Fe(NtKMSOO2]

Ferroin indicator:  dissolve 1.U85 g 1,10-phenantroline monohydrate and
      0.695 g ferrous sulfate, FeSO.,  7H20, in water and dilute to
      100 ml.  Alternatively, a commercially prepared indicator can
      be purchased.
Silver sulfate, AgaSOi*, reagent powder.
Mercuric sulfate, HgSOij, analytical-grade crystals.
Procedure
            Place several boiling stones or glass beads and 1.0 g
HgSOi, in a reflux flask.
            Transfer 0.5 to 2.0 g blended, wet sediment to the flask.
Wash the sediment into the sample flask with a minimum amount of
 * Corning 5000 or equivalent.
** Corning 2360, 915^8, or equivalent.
                                  3-393

-------
distilled water (25 ml).
            Add 25 ml 0.25 N K2Cr207 to the flask and mix thoroughly.
            Slowly, and with constant mixing, add 75 ml of sulfuric acid-
silver sulfate solution.  Ensure that the mixture is well mixed to avoid
localized superheating.
            Attach the sample flask to a condenser and reflux for 2 hr.
Should the added dichromate dissipate during reflux, either:  (l) repeat,
using a smaller sample size; or (2) carefully add additional 0.25 N.
K2Cr207 through the condenser.  Be sure to record any added dichromate.
            Allow the sample to cool and rinse the condenser with 1+0 to
50 ml distilled water.
            Add an additional 50 ml of distilled water to the sample
and allow to cool to room temperature.  Add 3 to 5 drops of ferroin
indicator and titrate with 0.25 N. Fe(NHit )2(SOi, )2 to a sharp color
change (blue-green to reddish-brown).
            For a blank, reflux 25 ml of distilled water, 25 ml 0.25 N
K2Cr207, 1 g HgSOij , several glass beads or boiling stones, and 75 ml
of sulfuric acid-silver sulfate solution for 2 hr.  Treat as a sample
and titrate with 0.25 N Fe(NH4 )2 (SO^ )2 after cooling and adding 3 to 5
drops of ferroin indicator.
Calculations
            The COD concentration of the sediment sample is calculated
as follows :
               COD mg/kg (wet weight) = (A " B) (N) (800)
                                               o
               COD mg/kg (dry weight) = (A -
where
        A = volume of 0.25 N_ Fe(NH4 )2(SOit )2 for blank titration, ml
        B = volume of 0.25 N. Fe(NHit )2 (SOi^ )2 for sample titration, ml
        N = normality of Fe(NH4 )2 (SO^ )z used for titration, eq/
     8000 = equivalent weight of oxygen, mg/eq.
        g = wet weight of sample, g
      % S = percent solids in sediment sample (expressed as a decimal
            fraction)

                                 3-391*

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                              References
1.  American Public Health Association.  Standard Methods for the
    Examination of Water and Wastewater Including Bottom Sediments and
    Sludges. lUth Edition.APHA; New York, New York.1193 p. (1975).
2.  Environmental Protection Agency.  "Manual of Methods for Chemical
    Analysis of Water and Wastes."  Environmental Monitoring and Support
    Laboratory, EPA; Cincinnati, Ohio (1979).
3.  Burns, E. R., and Marshall, C.  "Correction for Chloride Interference
    in the Chemical Oxygen Demand Test."  Journal of Water Pollution Con-
    trol Federation 37:1716-1721 (1965).
h.  Great Lakes Region Committee on Analytical Methods.   "Chemistry
    Laboratory Manual for Bottom Sediments."  U.  S.  Department of the
    Interior, Great Lakes Region; Chicago, Illinois.   96 p.  (1968).
                                 3-395

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                        SEDIMENT OXYGEN DEMAND

            Sediments are generally in a reduced chemical state and have
a potential to remove oxygen from overlying water.   This results from the
migration of dissolved oxygen to the sediment water interface followed
by subsequent chemical reaction and/or the migration of reduced chemical
species (ferrous iron, manganous manganese, sulfide) from the sediments
to the overlying water followed by subsequent oxidation.  The sediment
oxygen demand procedure characterizes sediments in  terms of rate of
exertion of oxygen demand.
            The procedure is a lengthly one that may require days or
weeks to acquire the data.   The actual time required will depend on the
rate of oxygen demand exertion by the sample.  A more restrictive aspect
of the sediment oxygen demand procedure is that it  should be run in situ.
                  i *
Edberg and Hofsten   compared in situ and laboratory-incubated sediments
and observed that laboratory-incubated samples only exerted 40 percent
of the in situ demand with a range of 9 to 100 percent.  The observed
rate of oxygen demand exertion doubled during 6 to  21 days of laboratory
storage and was attributed to the development of a  microbial population
and/or increased surface area exposure due to burrowing activity and
gas evolution.

                     Sample Collection and Storage

            The preferred method of running this procedure would be
in situ which would not require the collection of samples.  However, if it
is necessary to run the procedure in the laboratory, the sample should
be collected and placed in a glass or plastic container.  The container
should be completely filled to exclude any entrapped air and tightly
sealed.  The sediment oxygen demand procedure should be initiated
immediately on return to the laboratory.
            Only field moist samples should be used for the test as
dried and frozen samples will have been subjected to air oxidation.

* References for this section can be found on page  3-^03.
                                  3-396

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Sample handling and atmospheric contact with the moist sample should also
be minimized to decrease the effects of atmospheric oxidation.   During
transport, the samples should be kept at UC.

                  Procedures for Sediment Samplers1 '2

Method 1:  In Situ
Apparatus
Submerged chamber:  chambers have been reported in the literature
      as ranging from 33 to 115 cm in diameter.1'2   A schematic  repre-
      sentation of these chambers is shown in Figure 3-^7-   Chambers
      are embedded in sediments to an outside flange.  The flange
      prevents the apparatus from sinking further into the sediments
      and the chamber extension into the sediments restricts inter-
      stitial water transfer
      The apparatus should have an instrument port for inserting
      a dissolved oxygen electrode or a permanently mounted dissolved
      oxygen probe.  Additional sampling ports can be added if
      desired.  The chamber should also incorporate some method of
      stirring (magnetic stirrer, or a method of pumping water
      through the chamber) so that the dissolved oxygen measurements
      are not diffusion limited
Dissolved oxygen meter, Yellow Springs Instrument, or equivalent:
      equipped with sufficient cable to reach the sediments
Procedure
            Calibrate the dissolved oxygen meter.  Insert the electrode
in the chamber and lower the chamber in place.  Ideally, the chamber
should be inspected by divers to ensure the device has correctly pene-
trated the bottom sediments.
            Measure the dissolved oxygen concentration within the
chamber as a function of time.  Water temperature within the chamber
should also be measured and recorded.
            If additional parameters are to be measured, withdraw the
sample from the chamber with a syringe.  Process the water sample for
that specific parameter as indicated elsewhere in this manual.
Special precautions should be taken to avoid reaeration of the
sample.
                                  3-397

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            Outlet with
         i-Sliding Pipe
  Water  j   Tygon Tubing
Exchange
  Ports  1     ,  , ^
         I	Inlet
 Samp]ing
   Ports
                                                  Gas Vent Valves

                                                  Instrumentation
                                                       Ports
                                                           Flange
          A6 cm
                     K-
                           115 cm
                                              ^^^^dlment
                                              ^W \     Surface
                                               ->t
Figure 3-^7.    In situ sediment oxygen demand chamber
                         3-398

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Calculations
            The initial amount of oxygen in the chamber can be  calculated
as follows :
where
      2
     r  = radius of the chamber, cm
      h = internal height of the chamber above the sediments,  cm
      X = measured dissolved oxygen concentration, mg/&
     Ao = amount of oxygen in the chamber at time zero, mg
            The calculated oxygen values can -then be used to calculate
the rate of oxygen depletion:
                         R mg/m2/day =    ~ Al
                                       t2 - ti
where
                                     o
      R = rate of oxygen uptake,  mg/m /day
     Aa = oxygen within the chamber at time 2,  mg
     AI = oxygen within the chamber at time 1,  mg
     t2 = second sampling period, days
     tj = first sampling period,  days

Method 2:  Laboratory (SID)3
Apparatus
Laboratory oxygen analyzer
Magnetic stirrer with 1-in. Teflon-coated magnets
Incubator, 20C
Wide-mouth cylindrical jars:  with screw cap and sealed probe;  minimum
      mouth opening of 11 cm; height of 25 cm;  for use in making Oz
      uptake apparatus (Figure 3-^8)
Glass petri dish with cover
Glass petri dish support
Asbestos sheet, 15 by 15 by 0.5 cm
Reagents
Distilled water:  water used for solutions and for preparation  of the
      dilution water must be of the highest quality, distilled  from a
      block tin or all-glass still, contain less than 0.01 mg/& copper, and
      be free of chlorine, chloramines, caustic alkalinity, organic
                                  3-399

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      Dissolved
       Oxygen
       Probe
      (15cm  X
Asbestos  Shee
                                        Petri Dish  Cover
                                         and Cover  Remover
1 	
ith Jar

ng Bar 
OHI >,,/>,>.
^T
1 t" Q
f i
*


77777JJB^7T7T7} /ll|77^_r7.
0 _


	 Petr
Conta
-Pp t r '

.\>.^o^a
"" Magr
                                                     Oxygen
                                                      Meter
                                              Petri  Dish  Support
                                               Magnet i c  S t i rre r
      Figure 3-^8.    Laboratory sediment oxygen demand chamber

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material, or acids.
Phosphate buffer solution:  dissolve 8.5 g potassium dihydrogen phosphate,
      KH2POn; 21.75 g dipotassium hydrogen phosphate, KaHPOi* ;  33.^ g
      disodium hydrogen phosphate heptahydrate, NazHPOi*    7HzO; and
      1.75 g ammonium chloride, WHi^Cl, in about 500 ml distilled water
      and dilute to 1 .  The pH of this buffer should be  7.2  without
      further adjustment.  If dilution water is to be stored in the
      incubator , the phosphate buffer should be added just prior to
      using the dilution water.
Magnesium sulfate solution:  dissolve 22.5 g MgSOi*    THaO in distilled
      water and dilute to 1  .
Calcium chloride solution:  dissolve 27.5 g anhydrous CaCla  in distilled
      water and dilute to 1 .
Ferric chloride solution:  dissolve 0.25 g FeCls   6H20 in distilled
      water and dilute to 1 .
Procedure
            Store dilution water in cotton-plugged bottles for a
sufficient length of time to become saturated with dissolved oxygen.
            The water may also be aerated by shaking a partially
filled bottle or with a supply of clean compressed air.  Situations
may be encountered where it is desired to use stabilized river water
to check stream performance with laboratory procedure.  The distilled
water used should be as near 20G as possible and of the highest
purity.  Place the desired volume of distilled water in a suitable
bottle and add 1 ml each of phosphate buffer, magnesium sulfate,
calcium chloride, and ferric chloride solutions for each liter of
water .
            Set up the apparatus as shown in Figure 3-^-8.  Place two
asbestos sheets on a magnetic stirrer to prevent heat transfer and
place the wide-mouth jar on the asbestos sheets.
            Weigh a sub sample of moist, blended sediment and place in
a petri dish.  Cover the petri dish and place in the bottom of the
oxygen uptake chamber.  Support the petri dish off the bottom so the
magnetic stirrer can be used.
            Fill the uptake chamber with a known volume of dilution
water.  Remove the cover of the petri dish.
            Insert a standardized dissolved oxygen probe in the cap
of the oxygen uptake chamber and seal the chamber with the cap.  The

                                 3-U01

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cap should be lined with Teflon tape or plasticizer to ensure the seal  is
airtight.  The oxygen probe should be inserted far enough  so  that  it is
in the dilution water.
            Start the magnetic stirrer to simulate the flow in the
vicinity of the sediment source.  The agitation should not cause the
sediment to be resuspended.
            Take dissolved oxygen readings at various time intervals.
The number of readings will depend on the required frequency which,
in turn, will depend on the observed rate of oxygen uptake.
            The temperature in the uptake apparatus should also be
measured or, preferably, controlled in a constant-temperature room
as temperature can affect the rate of oxygen uptake.  The  rate of
oxygen uptake approximates Van't Hoffs' rule, with the rate approxi-
mately doubling for a 10-degree rise in temperature.
Calculations
            The initial amount of oxygen in the uptake chamber can be
calculated based on the volume of water used and the initial  oxygen
concentration as follows:
                             Ao =
where
     A  = amount of oxygen, mg
     V  = volume of water used, H
     C  = oxygen concentration at time zero,  mg/
            The rate of oxygen uptake can then be calculated based on
the change in the amount of oxygen in the chamber:
                                   A  -A
                             M = -r	-4	
where
      M = rate of oxygen uptake,  mg/g/day
     A^ = calculated amount of oxygen in the chamber at  time  1, mg
     Ap = calculated amount of oxygen in the chamber at  time  2, mg
     t  = elapsed time from the start of the test  to time  2,  days
     tp = elapsed time from the start of the test  to time  1,  days
      g = wet weight of sediment, g
                                 3-^02

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                              References
1.  Edberg, N., and Hofsten, B.  V.   "Oxygen Uptake of Bottom Sediments
    Studies in situ and in the Laboratory."  Water Research 7:1285-129^
    (1973).
2.  Sonzogni, W. C., Larsen, D.  P., Malueg, K.  W., and Schuldt,  M.  D.
    "Use of Large Submerged Chambers to Measure Sediment-Water Inter-
    actions."  Water Research Il;li6l-U61t (1977).

3.  Great Lakes Region Committee on Analytical  Methods.  "Chemistry
    Laboratory Manual for Bottom Sediments." U.  S. Department of the
    Interior, Great Lakes Region; Chicago,  Illinois.  96 p. (1968).
                                  3-U03

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 In  accordance with letter from DAEN-RDC,  DAEN-ASI dated
 22  July 1977, Subject:   Facsimile Catalog Cards for
 Laboratory Technical  Publications, a facsimile catalog
 card  in Library of Congress MARC format is reproduced
 below.
Plumb, Russell H., Jr.
  Procedures for handling and chemical analysis of
sediment and water samples / by Russell H. Plumb, Jr.
(Great Lakes Laboratory, State University College at
Buffalo). -- Vicksburg, Miss. :  U.S. Army Engineer
Waterways Experiment Station, 1981.
  478 p. in various pagings : ill. -- (Technical
report / U.S. Army Engineer Waterways Experiment
Station ; EPA/CE-81-1)
  "Prepared for U.S. Environmental Protection Agency/Corps
of Engineers Technical Committee on Criteria for Dredged
and Fill Material under Contract EPA-4805572010."
  "Monitored by Large Lakes Laboratory, U.S. Environmental
Protection Agency."
  Published in three-ring binder.
  Includes bibliographies.

  1. Chemistry, Analytic.  2. Dredging spoil
3. Fills (Earthwork).  4. Sediments (Geology).
Plumb, Russell H., Jr.
  Procedures for handling and chemical analysis : ... 1981.
                                                  (Card 2)

S. WaterAnalysis.  I. Environmental Protection
Agency/Corps of Engineers Technical Committee on
Criteria for Dredged and Fill Material.  II. United
States. Environmental Protection Agency. Large
Lakes Laboratory.  III. U.S. Army Engineer Waterways
Experiment Station.  Environmental Laboratory.  IV. Title
V. Series: Technical report (U.S. Army Engineer
Waterways Experiment Station) ; EPA/CE-81-1.
TA7.W34ep  no.EPA/CE-81-1

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