-------
Dissolved Oxygen Determination
7-6
-------
Dissolved Oxygen Determination
SCE*-
•Thermometer
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Figure 2. ELECTRODE ASSEMBLY FOR
DO DETERMINATION
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Figure 4. THE BECKMAN OXYGEN
SENSOR
OXYGEN ELECTRODE
L-Platinum Disk
Silver Ring
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Figure 3. DISSOLVED OXYGEN
ELECTRODE
GLASS
AMPLIFIER
mOlECTWI CASi
ELECTROLYTE
MEMMANE
Figure 5
7-7
-------
Dissolved Oxygen Determination
7 For convenience, the direct use of
parts per million in dissolved oxygen
measurements has often been employed.
Some workers regard percent saturation
as the more meaningful consideration.
8 In order to read the correct oxygen con-
centration accurately and continuously,
a fresh representative sample must be
maintained at the tip of the oxygen sen-
sor at all times during measurement.
Rate of oxygen diffusion is so slow that
it is unable to replenish even the minute
quantity consumed by the sensor.
9 Calibrating the oxygen sensor for ppm
dissolved oxygen measurements may be
accomplished by one of several methods:
a Grab sample method - correlation of
oxygen analyzer with Winkler method.
b Saturation technique - this involves
bubbling of air or oxygen through
water sample until saturation is
reached and then setting the oxygen
analyzed to read either ppm oxygen
or 100% saturation.
c Air calibration - since the partial
pressure of oxygen (160 mm) is the
same in air as it is in a 100% air
saturated liquid, the oxygen analyzer
can be calibrated simply by drying
off the tip of the oxygen sensor and
setting the instrument to read 100%
saturation.
B Yellow Springs Instrument Company Probe
Model 51 Oxygen Meter
1 Performance data, oxygen meter
(Figure 6)
a Range
0-25 ppm DO
0 - 50% DO saturation at 760 mm
b Accuracy
With direct calibration the error is
less than 1/2% saturation or 0. 25 ppm.
"O" RING
MFM8RANE
ANODE m
con IU-J-
The YSI Oxygen Probe
Figure 6
c Readability
0. 2% saturation or
0. 1 ppm
d Response time
90% of reading in 10 sec. dependent
on temperature and oxygen level.
e Temperature range of Og measurement.
5 to 45°C.
f Battery life
500 hrs.
2 Performance data, temperature probe
a Range 0 - 50°C
b Accuracy + 1. 5°C
c Readability 0. 2°C
3 Readings below surface of water:
a Instrument can be calibrated to read
directly in % saturation of Oxygen.
b If the measurement is to be made
below the surface of the water, the
pressure of the water must be taken
into consideration because the amount
of dissolved oxygen is related to the
total pressure of the layer of water
being analyzed.
c Automatic pressure - compensation
for sample depth can be effected by
calibration chamber design.
7-8
-------
Dissolved Oxygen Determination
4 Calibration
Whenever possible, calibration should
be performed at the same temperature
as the measurements (Figure 7).
a Assemble stopper (3) onto probe (4).
10
Figure 7
The Direct Calibration Method
b Insert probe-stopper assembly into
top of calibration chamber (5).
c Insert stopper (6) into bottom of
chamber (5).
d Chamber is filled with air, and con-
nected to tube (1). The pressure is
equalized in chamber at sample
depth. Oxygen in air is measured at
temperature of sample.
5 Measurement
a Transfer probe stopper from chamber
(5) to ring (7).
b Twist handle to induce flow past
membrane.
6 Theory of measurement of YSI oxygen
probe.
a The cathode is a gold ring imbedded
in a lucite block.
b The anode is a silver coil recessed
in the center well.
c The interior is filled with a solution
of KC1.
d A thin teflon membrane is stretched
^—across the end of the sensor.
^ff^"^ - '— «-S"«ai.,w
> e When a suitable polarizing voltage
{ is applied across the cell, oxygen
\ will react in the cathode causing a
current to flow through the cell.
C The Galvanic Cell DO Analyzer (Figure 8).
A lead anode-silver cathode galvanic
couple.
To Microommerer
Plastic Casting
Plastic Collar
Polyethylene
Membrane
Lens Paper
Lead Anode
Silver Cathode
THE GALVANIC CELL OXYGEN ANALYZER
Figure 8
7-9
-------
Dissolved Oxygen Determination
1 Lead is selected as the anode because
its electrode potential is sufficiently
negative to cause spontaneous oxygen
reduction without externally supplied
voltage.
2 KOH is the electrolyte.
a Residual current is small.
b Lead ions are soluble.
c A clean anode surface is maintained.
3 Electrode reaction
a Cathodic reaction
O2 + 2H2O + 4e ^ 4 OH"
b Anodic reaction
pb + 4OH" ^ pbC>2 + 2H2O + 2e
4 The salt effect
When the analyzer is used in water
samples, a correction factor may have
to be applied to the cell sensitivity.
a If the oxygen analyzer probe is cali-
brated in a salt solution identical
to the salt solution being investigated,
no correction factor is needed.
A correction need not be made for the
"salt effect" in waters having a
dissolved solid of less than 500 mg/1.
D Calibration Practice at SEC
In the water quality surveys, SEC workers
calibrate all DO probes daily, using the
Alsterberg Modification of the Winkler
procedure as the standard of comparison./
REFERENCES
1 A Dissolved Oxygen Primer. Beckman
Bulletin 7015. Beckman Instrument,
Inc. Fullerton, California. 1962.
2 Precision Scientific Co. Technical
Bulletin TS-68850. Precision Scienti-
fic Company. Chicago, Illinois.
3 Instructions for YSI Model 51 Oxygen
Meter. Yellow Springs Instrument
Company. Yellow Springs, Ohio.
4 Standard Methods for the Examination of
Water and Wastewater. APHA, AWWA,
WPCF. llth ed. 1960.
5 Porges, R. Dissolved Oxygen Determina-
tions for Field Surveys. WPCF 36,
1247. 1964.
7-10
-------
LABORATORY PROCEDURES
J
DISSOLVED OXYGEN
D. G. Ballinger*
I WINKLER METHOD - AZIDE MODIFICATION
A Reagents
1 Manganous sulfate solution - 364 g
MnSO4H2O in distilled water, filter,
dilute to 1 liter.
2 Alkaline-iodide-azide reagent - 700 g
KOH and 150 g KI in 950 ml distilled
water. Dissolve 10 g NaNs in 40 ml
distilled water. Add to alkaline-iodide
solution with constant stirring.
3 Sulfuric acid, concentrated
4 Starch indicator
5 Sodium thiosulfate solution - 0.025 N
B Procedure:
1 To a full sample bottle, with the tip of
each pipette below the surface of the
sample, add 2 ml manganous sulfate
solution and 2 ml alkaline-iodide-azide
reagent.
2 Replace the stopper, rinse under
running water and mix by inverting
4-5 times. Allow the precipitate to
settle. Repeat the inverting and
settling.
With the tip of the pipette
face of the sample, add 2
trated sulfuric acid.
=ibove/the sur-
Concen-
Stopper the bottle, rinse under running
water and mix to dissolve the precipi-
tate.
By means of a 200 ml volumetric flask,
properly modified, transferj!0_3jnl of
the treated sample to a 500 ml Erlen-
meyer flask.
6 Titrate with 0. 025 N THIO to a pale
straw color, add 2 ml starch indicator
and continue titrating to the disappear-
ance of the blue color.
C Calculation
Because of the addition of reagents, a por-
tion of the sample in the bottle has been
displaced. To compensate for this loss
of sample, the volume of treated sample
titrated must be adjusted as follows:
vol. of sample bottle
X 200
vol. of bottle - ml of reagents
= vol. of sample titrated
for the above procedure:
300 "
300 - 4
X 200 = 203 ml
Since the sulfuric acid displaced sample
which had been beoxygenated by previous
reaction, the 2 ml of acid does not enter
into the correction. When 203 ml of the treated
sample is titrated with 0. 025 N thio:
1 ml 0.025 N thio £ 1 mg/1 DO
II RIDEAL-STEWART MODIFICATION
A Reagents
1 Potassium permanganate solution -
6. 39 KMnO4 in 1 liter distilled water.
2 Potassium oxalate solution - 2 g
in 100 ml distilled water.
3 Potassium fluoride solution - 40 g
KF 2H2O in 100 ml distilled water.
4 Manganous sulfate solution - 364 g
MnSO4H2O in distilled water, filter,
dilute to 1 liter.
*In Charge, Chemistry, Technical Advisory and Investigations Section, DWSPC, SEC.
December 1965.
Reviewed
CH.O.do. 19.8.59
7-11
-------
Dissolved Oxygen Determination
5 Alkaline-iodide-azide reagent - 700 g
KO4 and 150 g KI in 950 ml distilled
water. Dissolve 10 g NaNs in 40 ml
distilled water, add to alkaline-iodide
solution with constant stirring.
6 Sulfuric acid, concentrated
7 Starch indicator
8 Sodium thiosulfate solution - 0.025 N
B Procedure
1 To a full sample bottle, with the tip of
each pipette below the surface of the
sample, add 0.7 ml concentrated sul-
furic acid, 2 ml potassium fluoride
solution, and 1 ml potassium permanga-
nate solution.
2 Replace the stopper, rinse under run-
ning water, and mix by inverting 4-5
times. If a violet color does not persist
after mixing, add more permanganate
solution in 1 ml increments until the
color persists for 5 minutes.
3 With the tip of the pipette below the
surface of the sample, add potassium
oxalate solution in 0. 5 ml increments
until the sample is free of permanga-
nate color. Note: place sample in dark
for a few minutes after each addition
of oxalate. Avoid excess oxalate by al-
lowing sufficient time for decolorization.
4 When the sample is free of permanganate
color, with the tip of each pipette below
the surface of the sample, add 2 ml man-
ganous sulfate solution and 3 ml alkaline-
iodide-azide reagent.If a precipitate
does not form, add an additional ml of
alkaline-iodide-azide reagent.
Replace the stopper, rinse under run-
ning water, and mix by inverting 4-5
times. Allow the precipitate to settle.
Repeat the inverting and settling.
With the tip of the pipette above the
surface of the sample, add 2 ml cone.
sulfuric acid.
7 Stopper the bottle, rinse under running -
water, and mix to dissolve the precipi-
tate.
8 By means of a 200 ml volumetric flask,
properly modified, transfer 206 ml of
the treated sample to a 500 ml Erlen-
meyer flask.
9 Titrate with 0.025 N thio to a pale
straw color, add 2 ml starch indicator
and continue titrating to the disap-
pearance of the blue color.
C Calculation
Because of the addition of reagents, part
of the sample in the bottle has been dis-
placed. To compensate for this loss of
sample, the volume of treated sample
titrated must be adjusted as follows
vol. of sample bottle
X 200
vol. of bottle - ml of reagents
= vol. of sample titrated
In the above procedure, a total of 9. 2 ml
of oxygen-bearing sample was displaced
by reagents.
300
300 - 9.2
X 200 = 206 ml
Note: If more than the prescribed amount
of any of the reagents is required, the
volume of treated sample for titration
must be adjusted.
When the correct volume of treated sample
is titrated with 0.025 N thio:
1 ml 0. 025 N thio % 1 mg/1 DO
III THERIAULT (SHORT WINKLER)
MODIFICATION
A Reagents
Same as for Azide modification
7-12
-------
Dissolved Oxygen Determination
B Procedure
The Theriault modification consists of the
Alsterberg modification, but performed
as rapidly as possible. The usual settling
of the precipitate is omitted and the
titration is performed immediately after
the sample has been acidified.
C Calculation
The calculation and reporting of results
for the Theriault modification are the
same as for the Winkler Method -
Azide modification.
7-13
-------
POLAROGRAPHIC ANALYSIS
D. G. Ballinger*
I INTRODUCTION
A Historical Background
The polarographic method was first de-
scribed by Heyrovsky, at Charles Universi-
ty in Prague, in 1922. Using the electro-
chemical laws postulated by Nernst Prof.
Heyrovsky developed the principles and
instrumentation for the current-voltage
relationships and called the method "po-
larography". Most of the concepts and
design factors have remained unchanged
to the present time.
The earliest polarographic instruments
were a recording type, using photographic
paper to trace the polarograms. The
electrode systems were nearly identical
to those in use today.
B Utility
The polarographic method, with its as-
sociated procedures, is one of the most
sensitive analytical techniques available.
It is widely used in research as well as
in routine analysis. Since the method is
relatively new, refinements in technique
and application are continually appearing.
II THEORETICAL BASIS
A Electrolysis
Polarography is defined as the electroly-
sis of a minute fraction of a solution
around a small easily polarizable elec-
trode. The electrolysis is confined to
that part of the solution which is in con-
tact with the surface of the small elec-
trode, while the body of the solution re-
mains unchanged. For purposes of this
discussion, a polarized electrode is one
which adopts the potential externally im-
posed on it, with no change in current.
The most familiar forms of electrolysis are
those involved in electroplating baths, or
in the production of gases at electrodes.
In all electrolysis, the chemical reactions
which occur at the electrodes are the re-
sult of the flow of electrons from the ex-
ternal source of voltage. In the galvanic
cell or battery, however, chemical re-
actions in the cell produce a flow of
electrons, which is current.
Consider the case of two platinum elec-
trodes suspended in a well-stirred solu-
tion of copper sulfate. When an increas-
ing external voltage is applied, no reaction
occurs until sufficient electrons are
available to reduce the copper ions;
Cu++
+ 2 e"
= Cu°
When this potential has been reached, the
copper ions will "plate out" on the cathode.
If current measurements are made and the
current-applied EMF is plotted, the curve
will appear as:
Current
Applied EMF *-
After the decomposition potential E^ is
reached, the reaction follows a straight
line, the slope being dependent on Ohm's
Law. This linear relationship will con-
tinue until all the copper ions in the solu-
tion are reduced, since the stirring will
bring all the ions to the electrode surface.
B Limiting Current
Now consider a similar case, where one
of the electrodes is very small and the
other large, and the solution is not stirred.
Under these conditions, the current will
*In Charge, Chemistry, Technical Advisory and Investigations Section, DWSPC, SEC.
CH.MET. 12a. 12.65
7-14
-------
Polarographic Analysis
rise until the concentration of Cu++ ions
at the surface of the small electrode
approaches zero. In such a quiescent
solution, ions can only reach the elec-
trode by diffusion. Further increase in
applied potential will not increase the
current linearly, since the current is
now limited by the supply of cupric ions.
In fact, the magnitude of the current will
depend upon the rate of diffusion of ions
to the electrode surface. A plot of the
current-voltage curve would appear as:
Current
Limiting
Current
E
When the concentration of ions at the
electrode surface approaches zero, the
concentration gradient across the inter-
face approaches a constant. The value
of this constant is a function of the con-
centration of ions in the body of the solu-
tion. Under such condition, the small
electrode is said to be "concentration
polarized, " i.e., the current flowing is
limited by the concentration gradient of
a specific ion at the electrode surface.
In the polarographic method the small
polarizable electrode is usually a mer-
cury drop falling from a capillary. The
capillary is selected to yield a new drop
every 3-5 seconds. With each new drop
a new electrode surface is provided,
eliminating all deposition products from
the previous electrolysis. Each drop is
an exact replicate of its predecessor,
making the electrode characteristic re-
producible.
C Quantitative Use
The limiting current is actually the sum
of three separate componentS:the residu-
al current, the migration current, and
the diffusion current. The residual cur- f
rent is the result of electrochemical re-
action at the surface of the small elec-
trode, independent of any specific ion
reaction. The residual current is nor-
mally very small and is proportional to
applied EMF. The migration current is
the result of electrostatic forces which
cause a flow of positive ions toward the
cathode and negative ions toward the
anode. The ions are said to "migrate"
toward the opposing electrode. If a large
excess of inert electrolyte is present in
the sample, the electrical resistance be-
tween the electrodes is very small and
the current is carried entirely by the ions
of the inert electrolyte. However, since
the electrons may reach the electrode
surface only by diffusion, the current
which flows is very small and is actually
dependent upon the rate of diffusion.
The diffusion current is the result of
diffusion of the ions into the electrode -
solution interface. The rate of diffusion
is controlled by the concentration gradient
between the ions at the surface of the
electrode and those in the body of the
solution (Fig. 1).
CONCENTRATION GRADIENT
AT D. M. E.
Figure 1
7-15
-------
Polarographic Analysis
Since the effect of the migration current
is negligible and the residual current is
constant at a specific applied EMF, the
diffusion current alone may be considered.
As indicated above, the diffusion current
is controlled by the concentration gradient,
which is in turn a function of the concentra-
tion of test ions. Measurement of the dif-
fusion current will yield a value directly
proportional to the concent ration in the body
of the solution. Thus the use of thepolaro-
graph as a quantitative instrument requires
the application of a definite potential and
the measurement of the resulting current.
Figure 2 shows a polarogram of copper
solution at four different concentrations.
Note that the wave height (current) is
directly proportional to the concentration
of copper in each solution.
In quantitative analysis with the polaro-
graph, the current-concentration relation-
ship is determined on a series of standard
of known concentration. A calibration
curve is constructed and a proportionality
factor is calculated, using the equation:
C = K (D -
where:
C = concentration
K = proportionality factor
D = Diffusion current at
C concentration
D = Diffusion current at
o
O concentration
For subsequent determinations of unknown
samples, the diffusion current is measur-
ed at the same applied EMF and under the
same conditions and the concentration
calculated from the proportionality.
WAVE HEIGHT AT VARIOUS CONCENTRATIONS
RESIDUAL CURRENT
O.I
0.2 0.3 0.4 0.5 0.6 0.7
Figure 2. NEGATIVE POTENTIAL
VOLTS VS. SCE
7-16
-------
Polarographic Analysis
D Qualitative Use
The decomposition potential E^, at which
electrode reaction begins, is effected by
the concentration of reducible ions. How-
ever the half-wave potential, (the potential
at which the current is equal to one-half
of its limiting value) is independent of con-
centration and specific for the particular ion
being reduced. It may therefore be used for
qualitative determination. Et can be de-
termined graphically, as shown in Fig. 3.
Tables of half-wave potentials are avail-
able in the literature and are helpful in
the identification of unknown ions. From
these data, the composition of a mixture
of several ions may be determined. The
polarogram of such as mixture is shown
in Fig. 4. Since the supporting electrolyte
influences the half-wave potential, the
proper electrolyte must be specified.
Ill INSTRUMENTATION
A Circuitry
As shown in the circuit diagram, Fig. 5,
the polarographic instrument has:
1 a source of EMF, generally 0-3 volts
2 a voltameter for measuring the EMF
3 a variable resistance for controlling
the applied EMF
4 a sensitive galvanometer for measur-
ing the resultant current
5 a cell, consisting of two special elec-
trodes.
GRAPHIC DETERMINATION
OF HALF-WAVE POTENTIAL
0.
0.2 0.3 0.4 0.5 0.6 0.7 0.8
Figure 3. NEGATIVE POTENTIAL
VOLTS, VS. SCE
09
1.0
7-17
-------
Polarographic Analysis
Most instruments also incorporate ad-
ditional circuits for reversing polarity,
selecting galvanometer range, or auto-
matically recording the current-voltage
curves. Regardless of the complexity
or cost of polarographic instruments have
the five basic components shown above.
B Cell
#
Figure 6 illustrates the electrode as-
se'mbly normally used in polarographic
work. The mercury level, which must
be held constant, is controlled by the
leveling bulb. The cell contains the
^ POLAROGRAM OF MIXTURE
NEGATIVE POTENTIAL
IP VOLTS VS. SCE
0.5 0.6 0.7 0.8 0.9 1.0 1.1 1.2 1.3 1.4 1.5 1.6
7-18
-------
Polarographic Analysis
B ~^-::-
TYPICAL POLAROGRAPHIC CIRCUIT
Figure 5
dropping mercury electrode and a reference
electrode, generally saturated calomel,
connected by means of a salt bridge.
C Variables Influencing the Diffusion
Current
A study of the relationship of cell design
to current produced has shown that the
current is influenced by several variables:
1 Size of the mercury drop- the amount
of current flowing is proportional to
the surface area of the drop. Therefore,
drop size must be constant for quanti-
tative work. Changes in capillary size
or length or in static pressure on the
mercury will influence the size of the
drop.
2 Drop time - For reproducible results
the life of each drop should be 3-5
seconds. Shorter drop time will cause
stirring action around the electrode;
longer time will permit interference
from vibration and adsorption of de-
composition products. Drop time is
affected by capillary characteristics,
electrolyte composition and concentra-
tion, applied potential, and pressure on
the mercury.
3 Temperature - Changes in tempera-
ture affect the drop time and size of
the mercury drop, the migration cur-
rent, the rate of diffusion, and the
adsorption of decomposition products.
Normally the cell and sample are im-
mersed in a thermostated water bath
to control temperature fluctuations.
Where the sample must be analyzed
without temperature control, a small
thermometer is incorporated in the
electrode assembly and suitable tem-
perature correction calculated. Tem-
perature coefficients have been found
to be approximately 1. 5% change in
current for each 1° change in tem-
perature.
4 Oxygen Interference - For most polaro-
graphic determinations it is necessary
to remove dissolved oxygen from the
sample. Oxygen produces a very
prominent polarographic wave over
the region 0.1 - 1.8 volts vs. SCE.
Removal of the oxygen is accomplish-
ed by the addition of alkaline sodium
sulfite solution or by bubbling an inert
gas such as nitrogen through the sam-
ple.
5 Reference Electrode - In order to
accurately control the applied potential
a reference electrode must form one
of the half-cells in the electrode as-
sembly. Generally a saturated calomel
electrode is used. To prevent a shift
in the half-wave potential the SCE
should be large and the salt bridge
must have a low resistance junction.
E Commercially Available Instruments
1 Manual Instruments - requiring the
manual control of the applied voltage
through a rheostat. These are non-
recording instruments, so that polaro-
grams are developed manually from
galvanometer readings.
Sargent Model III - Uses 1. 5 v dry
cells as a source of EMF. Current is
indicated on a 300 mm. curved scale.
A moving coil galvanometer measures
current over a wide range. This instru-
ment is versatile, yet has accuracy
comparable to more expensive models.
It is suited to routine laboratory analysis
7-19
-------
Polarographic Analysis
Wire
Leveling
Bulb
Glass Bulb
Mercury
Level
Plastic Tubm
Gloss Tubing
Plastic Tubing
--Wire
ELECTRODE ASSEMBLY
Rubber
Cap
Glass
Saturated
Colonel Electrode
Capillary
Glass
Sample
Figure 6
7-20
-------
Polarographic Analysis
and may be adapted to rotating elec-
trodes. . .. $475.
Fisher Electropode - A compact, self-
contained manual instrument. The cell
and electrode assembly are mounted on
the front of the cabinet, but may be dis-
mounted for use in a water bath or
special cell. The galvanometer as-
sembly is rugged enough to make the
instrument portable. . . . $615.
Recording Instruments - these auto-
matically apply an increasing potential
at a constant rate. The movement of
the chart recorder is synchronized
with the voltage divider so that the cur-
rent - voltage curves are correctly
plotted.
Sargent Model XXI - a pen recording
instrument, providing maximum ver-
satility. The polarogram is recorded
on roll type chart. A good instrument
for research in electrochemical re-
actions. ...$2580.
Sargent Model XV - a good instrument,
having all the necessary features at a
reasonable price. This polarograph
can be adapted to the use of the Micro-
range Extender (Sargent) yielding sen-
sitivities to 0. 0001 ua/mm. . . . $1585.
REFERENCES
1 Kolthoff, I. M., and Lingane, J. J. Polaro-
graphy. Interscience Publishers. New
York. 1952.
2 Muller, O. H. The Polarographic Method
of Analysis. Journal of Chemical
Education. Eaton, Pa. 1941.
3 Meites, Louis. Polarographic Techniques.
Interscience Publishers. New York.
1955.
4 Milner, G. W. C. The Principles and
Applications of Polarography and Other
Electroanalytical Processes. 1957.
7-21
-------
S_r> /
& py &)
POLAROGRAPfflC DETERMINATION OF DISSOLVED OXYGEN FOR BOD
N. C. Malof*
I EQUIPMENT
A Recording Polarograph, /vith dropping
mercury electrode (approximately 3 sec-
onds) and saturated calomel electrode.
B Sample bottle, 125 ml glass stoppered.
II REAGENTS
Electrolyte - dissolve 0. 375 g methyl red
and 0. 2 g KOH in 50 ml distilled water. Add
18. 75 g KC1 and dilute to 100 ml with distilled
water.
Ill SAMPLE PROCEDURE
A Instrument
1 Turn instrument on and allow approx-
imately 20 minute warm-up.
2 Check instrument (daily) as described
in Part C.
3 Check sample temperature. Should be
19-2l°C.
4 Add 0. 5 ml electrolyte, below surface
of sample. Stopper with rubber stopper
and mix.
5 Set instrument:
Sensitivity 0. 020 Range 0 to -1 volts
6 At 0 Applied voltage, adjust recorder
pen to 10 mm line on chart with dis-
placement control.
7 Set Applied voltage to 40%. Allow pen
deflection to stabilize until peak heights
are equal.
8 Set Applied voltage to 0.
9 Measure total mm peak height and
subtract 10 mm.
10 Using calibration table, calculate DO
in mg/1. Repeat steps 3 to 10 for
each sample.
B Standard Curve and Calibration Table
1 Prepare a series of at least 10 samples
having a range of DO values from
1-9 mg/1 by bubbling nitrogen gas
' ' through tap water.
2 Fill duplicate bottles, using 125 ml gas
stoppered bottle for polarographic
determination and standard BOD bottle
for Winkler method.
3 Determine peak height on each standard,
as in steps 3-8 in Sample Procedure.
4 Determine DO by Winkler method on
corresponding bottles.
5 Plot peak height vs Winkler DO on
linear graph paper.
6 Using method of least squares, find
equation for line of best fit.
7 Using equation of best fit, prepare a
table for converting mm peak height to
mg/1 DO. This table is only valid for
the electrodes and mercury height used,
and for the sample temperature at
which the peak heights were measured.
C Daily Instrument Check
1 After approximately 20 minutes warm-
up, turn Operation control to Release
Chart position.
2 Check standardization by holding
"Standardization" control first to Volt-
age, then to Current, observing pen
travel. Pen should move slowly to left
on Voltage and on Current. If necessary,
adjust pen travel with appropriate
controls.
#Chemist, Technical Advisory and Investigations Section, DWSPC, SEC.
CH.O.do.27a. 12.65 7-22
-------
Polarographic Determination of DO jor BOD
3 Turn Cell control to Rs (standard
resistor) .
4 Set Voltage Range to 0 to - 1 volts,
Voltage Applied toj) and sensitivity to
0.020.
5 Adjust pen to 10 mm line on chart with
displacement control.
6 Turn Operation control to Record and
place pen on paper.
7 Turn Voltage Drive control to 0 to 100%.
8 Pen should trace a straight line, sloping
sharply to left.
For sample, adjust mercury height to
mark, check to see that mercury is
flowing in regular drops. Turn Cell
control to Normal Polarity.
IV LABORATORY EXERCISE FOR
BOD PROCEDURE
Place 1 liter of effluent into 4 liter bottles,
add 3 liters of dilution water (25% dilution).
Mix and aerate with diffuser and syphon
into 10 large (300 ml) DO bottles and 5
small (125 ml) bottles.
Run initial Winkler DO on large DO bottles
and incubate the other 8 bottles. Run initial
polarograph DO on 1 small bottle and incubate
the other 4 bottles. Run Winkler and polaro-
graph DO according to the following schedule:
Plot Summation (S) of each Winkler DO
determination (ADO) obtaining a 7-day BOD
curve. Determine k rate by the Moment's
method using a time (t) of 7 days. Repeat
plot and calculations for polarograph DO
determinations.
DAYS
Initial Wed. Thurs. Fri. Sat. Sun. Mon. Tues.
Day
Bottles
iWinkler
Bottles
Polarograph
0
2
1
2
2
2
3456
2222
7
2
7-23
-------
DISSOLVED OXYGEN PROBE LABORATORY
N. C. Malof*
I CALIBRATION OF DISSOLVED
OXYGEN PROBE
A Precision Scientific
1 Prepare duplicate DO samples and run
one of them by the Winkler Method.
2 Place a stirring bar in other bottle and
set it on magnetic stirrer.
3 Check thermistor by turning large knob
on meter to "temperature adjustment".
Turn needle to 40°C on meter with
smaller "temperature adjustment" knob.
4 Measure temperature of sample with
thermistor.
5 Remove thermistor from bottle and
insert probe. Turn on magnetic stirrer
to 1/2 or 3/4 speed being careful not to
cause air bubbles.
6 Turn knob to "oxygen" and read "ua" on
©2 scale.
7 Divide "ua" by Winkler DO value and
this gives the sensitivity coefficient
ua
^ DO
8 To determine DO of unknown sample,
divide ua by .
D0 = f
This meter is not temperature com-
pensated and does not read direct DO in
mg/1. A new sensitivity coefficient (4>)
must be calculated for each temperature
and calibration should be checked every
day.
B Yellow Springs, Inc.
1 Prepare duplicate DO samples and run
one of them by the Winkler Method.
2 Place a stirring bar in other bottle and
set it on a magnetic stirrer.
3 Check thermistor by turning knob to
read "red line" and then using adjust-
ing screw, set needle to red line on
meter face.
4 Measure temperature of sample with
thermistor.
5 Remove thermistor from bottle and
insert probe. Turn on magnetic stirrer
to 1/2 or 3/4 speed being careful not to
cause air bubbles.
6 Turn knob to "read" and adjust meter
to same DO value as Winkler. The
meter is now direct reading in mg/1
DO.
This meter is not temperature com-
pensated but does read directly in mg/1
DO. A temperature factor must be
applied at other temperatures.
The calibration of this probe should be
checked daily.
C Weston and Stack, Inc.
1 Prepare duplicate DO samples and run
one of them by the Winkler Method.
2 Check the thermistor by turning knob
on front of meter to "temperature". Push
"temperature check" button on left
side of case and needle should read
50°C. If not, turn "temperature adjust-
ment" screw on right side of case.
*Chemist, Technical Advisory and Investigations Section, DWSPC, SEC.
CH.O. do.lab. 2. 12.65
7-24
-------
Dissolved Oxygen Probe Laboratory
Insert probe into bottle making sure that
micro-switch on side of probe closes
and that vibrator on probe is moving.
Read temperature by turning knob to
"temperature".
Turn knob to "DO 1-mult"1" and set
needle to read same value as Winkler
DO. The meter is now direct reading
in mg/1 DO at any temperature.
This meter is temperature compensated
and will read directly in mg/1 DO. A
calibration check should be performed
daily.
REFERENCES
J Precision Scientific Co. Technical Bulletin
TS-68850. Precision Scientific Compan:
Chicago, Illinois.
2 Instructions for Y.S.I. Model 51 Oxygen
Meter. Yellow Springs Instrument
Company, Yellow Springs, Ohio.
3 Instruction for Weston and Stack Model
300 Dissolved Oxygen Probe. Weston
and Stack, Inc., Newtown Square,
Pennsylvania.
7-25
-------
, 3
-?-
3. ij
// c^
/32_
7.9
.)
^TRfc "^'S U<*.
-c- 3
-------
BOD TEST PROCEDURES
D.G. Ballinger*
I DIRECT METHOD
A Application
With relatively clean surface waters the
BOD may be determined by incubation of
the undiluted sample for the prescribed
time interval. This method is applicable
only to those waters whose BOD is less
than 8 mg/1. If the waste is known to be
lacking in proper biota the seeding pro-
cedure in III should be used.
B Procedure
1 Fill three 300 ml glass stoppered bottles
with the sample, making sure that no
air bubbles are entrapped and that the
bottles are filled to overflowing before
inserting the stoppers.
2 Determine the DO concentration on one
of the bottles by the appropriate Winkler
modification. This concentration is
reported as "initial DO".
3 Incubate the two remaining bottles at
20 C in complete darkness. The in-
cubated bottles should be water-sealed
by inversion in a tray of water or by
using a special water-seal bottle.
4 After 5 days of incubation, determine
the DO on the two bottles. Average
the DO concentration of the duplicates
and report as "Final DO".
5 Calculation
Initial DO - Final DO = 5 day BOD in
mg/1.
II DILUTION METHOD - UNSEEDED
A Application
Where the BOD of the sample is greater
than 8 mg/1, the sample must be diluted
to a concentration which will yield a DO
depletion less than that amount in 5 days.
Suggested dilutions are shown in the table
below:
If the waste is known to be lacking in
proper biota the seeding procedure in III
should be used.
B Procedure - Bottle Dilution Technique
1 Using an assumed or estimated BOD
value as a guide, calculate the appro-
priate factors for a range of dilutions
to cover the desired depletions. De-
j3j£JaonJutxj^^Apfie,»LiQ. -. 90% qf the
initial DO will give the most reliable
results.
____
less than J^mg/1 or_inore,thanm8^mg/l
may not be reliable. At leasf three ^^
dilutions in duplicate should be used.
Accurately measure the required amounts
of sample into 300 mlglass stoppered bottles.
Type of Waste
Strong Industrial Waste
Normal Sewage
Treated Effluents
Polluted Surface Waters
Estimated 5 day BOD
500 - 5000
100 - 500
20 - 100
5-20
Dilution
0.1- 1%
1 - 5%
5 - 25%
25 - 100%
*In Charge, Chemistry, Technical Advisory & Investigations Section, DWS&PC, SEC.
December 1965.
Reviewed
CH. O. bod. 43a. 8.63
3-1
-------
BOD: Test Procedures
Fill the bottles completely with dilution
water (Standard Methods l*£h Edition
page &c&T. \ «"
Using the appropriate Winkler modifi-
cation, separately determine the DO
concentration of the waste and the di-
lution water and calculate the "initial
DO". If the waste represents 1% or
less of the total volume, or it is known
to have a DO of practically zero, the
calculation should be based on the DO
of the dilution water.
Incubate the bottles at 20°C in complete
darkness. The incubated bottles should
be water-sealed by inversion in a tray
of water or by using a special water-
seal bottle.
After 5 days of incubation determine the
DO on the bottles. Average the DO con-
centration of the duplicates and report
as "Final DO".
7 Calculation:
(Initial DO - Final DO) X
300
ml waste per bottle
= 5 day BOD in mg/1
C Procedure - Cylinder Dilution Technique
1 Using an assumed or estimated BOD
value as a guide, calculate the factors
for a range of dilutions to cover the
desired depletions. Depletion in the
range of 40 - 90% of the initial DO will
give the most reliable results. Dilutions
showing final DO less than 1 mg/1 or
more than 8 mg/1 may not be reliable.
At least three dilutions in duplicate
should be used.
2 Into a one liter graduated cylinder (or
other similar container) measure ac-
curately the required amount of sample
to give one liter of diluted waste. Fill
to one liter mark with dilution water
(Standard Methods, 'ilth Edition, page
319). Carefully mix by stirring, avoid-
ing the entrapment of air bubbles.
3 Siphon the mixture from the cylinder
into three 300 ml glass stoppered bot-
tles, filling the bottles to overflowing.
4 Determine the DO concentration on one
of the bottles by the appropriate Winkler
modification and record as "Initial DO".
5 Incubate the two remaining bottles at
20°C in complete darkness. The incu-
bated bottles should be water-sealed by
inversion in a tray or by using a special
water-seal bottle.
6 After 5 days of incubation, determine
the DO on the bottles. Average the DO
concentration of the duplicates and re-
port as "Final DO".
7 Calculation
(Initial DO - Final DO) X
1000
ml sample used
= 5 day BOD in mg/1
III DILUTION METHOD - SEEDED
A Application
Many industrial wastes are sterile, due to
their chemical composition or the manu-
facturing process involved. These wastes
must be seeded with the proper type and
number of organisms to obtain correct
BOD values. The seed may be stale do-
mestic sewage, stream water, or an accli-
mated culture complex.
B Procedure
1 Calculate the percentage of seed re-
quired to produce at least 0. 6 mg/1
5 day BOD.
2 Calculate the proper waste dilutions as
in II B or II C above. Reduce the con-
centration of waste sufficiently to allow
for the seed depletion.
3 Measure the required amount of waste
as directed in II B or II C.
-------
BOD: Test Procedures
\ Add approximately half of the required
amount of dilution water to the sample.
This is necessary to assure that the
concentrated waste does not exert a
toxic effect on the seed organisms.
5 Measure the calculated amount of seed
into the bottle or cylinder and fill with
dilution water. Siphon into bottles if
using the method in II C.
6 Calculate (n-B-4) or determine (II-C-4)
the initial DO.
7 Incubate the bottles at 20°C in complete
darkness. The incubated bottles should
be water-sealed by inversion in a tray
of water or by using a special water-seal
bottle.
8 After 5 days of incubation, determine
the DO on the bottles. Average the DO
concentration of the duplicates and re-
port as "Final DO".
9 Calculations
/
Seed Correction
The value applied as a seed correction
is obtained by determining the BOD of
the seed itself. Either the Direct Method
(I-B) or the Dilution Method (II-B or H-C)
may be used, depending upon the strength
of the seed material. It is essential that
enough seed is used to produce normal
demand. Generally, a depletion of 3-6
mg/1 in 5 days indicates satisfactory de-
oxygenation. The following formula is
used for the seed correction:
Depletion in
seed correction X
bottle
%seed added to sample
% seed used for seed
correction deter.
= Seed Correction
BOD Calculation
(Initial DO - Final DO - Seed Correction)"
X Dilution Factor = BOD, - of the Waste
Example:
Data
Waste dilution used 2%
Seed cone, used 0.4%
Seed correction cone. 3%
Initial DO 9.0%
Final DO 3.0
Seed correction depletion 6. 8
Seed Correction
6.8X^1 = 0.9 mg/1
O
BOD Calculation
K9.0 - 3.0) - 0.9JX 50
= 255 mg/1 5 day BOD
IV IMMEDIATE OXYGEN DEMAND
A Application
In certain industrial wastes reducing agents
such as sulfites, sulfides, or ferrous iron
will cause an immediate chemical demand
on the DO. In order to arrive at the true
biochemical demand it is necessary to
differentiate between the two demands.
B Procedure
1 Prepare additional duplicate bottles of
each of the dilutions used in the standard
BOD test.
2 Determine the initial DO as directed
above.
3 Incubate the immediate demand bottles
for 15 minutes.
4 Determine the final DO on these bottles
and calculate the 15 minute depletion.
Record as "Immediate Oxygen Demand".
8-3
-------
BOD: Test Procedures
5 Calculation
Total Demand - Immediate Demand
= 5 day BOD
V PRECISION OF THE TEST
Precision of the BOD Test depends upon the
precision of the DO determinations involved
and the reproducibility of the biological activi-
ty in the individual bottles. Standard deviation
of the test has been calculated as 0.07 - 0. 11
ml of oxygen demand titrated. That is, the
individual BOD value is correct to approxi-
mately
0.09
— Decimal Fraction of Sample Used
VI REAERATION METHODS
A Application
Because the dilution method does not re-
produce the waste concentrations which are
found under natural conditions, some inves-
tigators advocate incubation of the undiluted
sample with periodic reaeration when de-
pletion has reduced the DO to approximately
1.0 mg/1.
B Procedures
1 Elmore Method
A relatively large volume of the sample
is stored in an unsealed bottle. Small
bottles are withdrawn in sets of 5 or
more, sealed, incubated, and the DO
determined at appropriate intervals.
When the DO concentration in the
smaller bottles reaches 1.0 mg/1 a
new set is withdrawn from the large
unsealed bottle.
2 Orford Method
The deoxygenation is carried out in a
large sealed jug from which samples
for DO are withdrawn at appropriate
intervals. To maintain the waste level
and a sufficient DO in the jug, additional
waste is added from a second open
container.
C Advantages and Limitations
1 All dilution water problems are elimi-
nated in these methods.
2 The limitations imposed by the limited
solubility of oxygen are removed.
3 The test is generally more representative
of stream conditions.
4 Toxicity is not readily apparent, and
may escape detection.
5 In spite of the greater range, the methods
are restricted to rather low BOD (<50
mg/1 5 day BOD).
5-4
-------
BOD: Test Procedures
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-------
BOD: Test Procedures
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Final
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Reading
Initial
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Actual
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oi
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8-6
-------
EFFECT OF SOME VARIABLES ON BOD
D. G. Ballinger and J. W. Mandia*
I TIME
A Importance
An examination of the basic BOD equation,
y = L (1__- 10~K"i), and the resultant ex-
ponential curve, will emphasize the im-
portance of time in the oxidation reaction.
The amount of oxidizable material react-
ing at time "t" depends upon the time in-
terval between the beginning of the reaction
and time "t". Obviously, the longer the
elapsed time, the more nearly complete
the reaction, but the percentage of avail-
able material involved in unit time is con-
stant throughout the whole period of
oxidation.
B 5 Day BOD
In the interest of standardization of the
test procedure, the 5 day interval has been
selected as the elapsed time. At 5 days
the initial lag period often encountered has
been passed and the reaction has attained
its normal rate. However, the oxygen
demand at 5 days is only a part of the total
demand of the waste material; it is mere-
ly a reference point on the oxidation curve.
-L iTC VcLLUG ODTSLltlGQ £Lt 3 Cl3.yS IjQUSX uG 1H""
terpreted in light of the velocity constant
C Determination of Constants
When y values are obtained at equal time
intervals during the oxidation reaction,
these values may be used in the determi-
nation of the constant "k" and "L". To
insure accurate demand values, it is es-
sential that the time intervals be carefully
controlled. This is especially true of the
early periods, when a few hours may pro-
duce large changes in demand.
D Complete Oxidation
Theoretically, the time required for com-
plete satisfaction of the BOD is infinitely
long.
-IH-
y = L(l - 10 Kt)
when y = L, the quantity 1-10
-kt
therefore: 10 =0 and t = °°
= 1
The practical time for complete oxidation
can be determined experimentally. For_
domestic sewage morethan 100 days is
necessary; for some industrial waste ma-
terials a much longer period may be
required.
II TEMPERATURE
A Effect on Oxidation Rate
Temperature is one of the important con-
trolling factors in any biological system.
In the BOD reaction, changes in tempera-
ture produce acceleration or depression
of the rate of oxidation. Figure 1 shows
the changes in the value of k at temper-
atures from 0 - 25°C.
B Test Temperature
In the BOD test procedure an arbitrary
temperature must be selected, in order
that the test may be reproducible. Though
a wide temperature range exists under
natural conditions, 20°C has been chosen
as representing the median temperature.
Incubation of the test containers at 20°C
for the whole period is now accepted
practice.
C Temperature Correction
When it is necessary to calculate the rate
of oxidation at a temperature other than
20 , the following relationship may be
used:
- T2>
*In Charge, Chemistry, Technical Advisory & Investigations Section, DWS&PC, SEC, and
Chemist, DWS&PC Training Activities, SEC. Reviewed December 1965.
CH. O.bod. 42a. 8.65
3-7
-------
BOD: Effect of Variables
5 M 15 30 35
DEGREES C
'k WITH CHANGE IN TEMPERATURE
where;
k = velocity constant at temperature T-^
k0 = velocity constant at temperature T0
& &
9 = temperature coefficient, for which
Streeter and Phelps obtained the
value 1.047.
HI pH
A Effect on BOD
The organisms which accomplish the bio-
chemical oxidation of organic matter are
acclimated to a narrow pH range. The
normal range for these organisms is pH
^ ,' 6.5 - 8. 3. Outside this range, the rate
oif ISidation is depressed. Figure 2 illus-
trates the 5 day BOD values obtained from
pH 4. 5 - 9.0.
B Adjustment of Concentrated Samples
When wastes are more acid than pH 6, 5
or more alkaline than pH 8. 3, adjustment
to pH 7. 2 is necessary before reliable BOD
1 values can be obtained.
C Dilution Samples
In the dilution technique, the Formula C
JL„ dilution water is buffered at pH 7. 2. To
' insure proper pH control, the sample-di-
lution waterrmixfure"should be checked.
IV ESSENTIAL MINERAL NUTRIENTS
A Importance
In 1932 Butterfield investigated the role of
certain minerals in the biochemical oxida-
tion of sewage and concluded that a balanced
pH 8
4 56 7 8 9 10
Time in Days
Effect of Mineral Nutrients on BOD
-------
BOD: Effect of Variables
mineral substrate was necessary for normal
oxidation. In addition, he found that the lack
of adequate nitrogen and phosphorus pro-
duced low BOD values. (Figure 3)
With some industrial wastes, the addition
of nitrogen and phosphorus significantly
' increases the efficiency of BOD satisfaction.
B Standard Methods Dilution Water
The dj.iu,ti.p.a
test is designed to grpYide^ the essential
mineral nutrients (including N and P) in
approximately the concentrations found in
rialuraTwafer.
V MICROBIOLOGICAL POPULATION
A Need for Complex Flora and Fauna
The work of Butterfield, Purdy, and
Theriault in 1931 demonstrated that the
biochemical oxidation process is accom-
plished by a complex action involving bio-
logical forms. Their data is summarized
by the graph in Figure 4. Note that the
highest BOD values were obtained when the
normal mixed population was used. Later
studies have emphasized the need for an
adequate mixed biota in BOD evaluation.
B Acclimatized Organisms
Early investigations into the mechanism
of the BOD dealt primarily with domestic
sewage. In this waste the organisms neces-
sary for oxidation are present initially in
adequate numbers and variety. Streams
receiving domestic wastes are populated
by biological forms which are able to readi-
ly utilize the organic matter as food.
With the expansion of industry in the past
20 years, new and complex waste materials
were added to the receiving streams, cre-
ating new environmental and nutritional
conditions. In most cases the receiving
streams have adapted to the new materials
through the development of acclimated popu-
lations. With certain exceptions, the or-
ganisms which exist in the stream at some
distance below a particular industrial out-
fall are those which are acclimated to the
waste and can satisfactorily accomplish
the biochemical oxidation of that material.
C Seeding
In the laboratory determination of the BOD,
it is ess^nEariHaFWe "prgaijisiiii, present
in the sample bottle during,.incubation be
similar in type and numbers to those which
will oxidize the waste unter natural con-
ditions. As indicated above, dome Silt
sewage and surface waters contain the
proper biota. Many industrial wastes, how-
ever, are lacking in biological activity
initially and the samples must be "seeded"
with organisms before incubation in the
BOD test. The selection of the proper
seed is a prime requisite for accurate BOD
evaluation.
All forms in river water
Mixed bacteria & plankton
Pure culture B. Aerogenes
Mixed culture bacteria
Pure culture B Aerogenes
5 6 7
Time in Days
Effect of Biological Forms on Oxygen Depletion
Figure 4
-------
BOD: Effect of Variables
D Sources and Types of Seed
Stale Sewage
For waste samples having organic corny
position similar to sewage, the most
reliable seedis domestic sewage which
has been stored at 20°C for approximate-
ly 24 hours. During the storage period
the organisms typical of fresh sewage
have been supercededby a population
representative of the oxidation stage. In
order to maintain an aerobic condition,
it is sometimes necessary to aerate the
sewage during the storage period.
2 Stream Water
The adaption of receiving water to in-
dustrial effluents has been mentioned
in C above. Because of the acclimated
organisms present below the outfall,
such streams offer an excellent source
of seed for the BOD determination. With
few exceptions, carefully selected seed
from the receiving stream will yield the
highest BOD values. When stream water
is used for seed, nitrification difficulties
(Vn) may increase.
3 Special Seed
Occasionally, because of the exotic nature
of the industrial waste, it is necessary
to artificially develop a culture complex
which will oxidize the waste material.
This is done by starting with a large va-
riety of organisms (as in sewage) and
feeding the culture with gradually in-
creasing amounts of the waste until an
acclimatization process has produced
organisms especially adapted to utiliza-
tion of the waste components. Such a
culture is obviously an excellent seed
for use in BOD tests on the particular
waste.
In waste treatment facilities, the same
procedure is followed and similar accli-
matization occurs. Effluents from such
installations can also be used as seed in
the BOD test.
E Quantity of Seed Required
The amount of seed required to produce
a normal rate of oxidation must be deter-
mined experimentally. The most frequent
error is the use of insufficient seed.
Figure 5 illustrates the work of Ludzack
in the determination of satisfactory seed
concentrations. When 0.1% seed was
used a definite lag period occurred. In-
creasing the seed concentration to 0. 2%
eliminated the lag. Further increase in
seed did not increase the oxidation rate.
Obviously the concentrations used in this
study would not necessarily apply to other
wastes and other seed types.
Calculation of the ratio of the 2-day and
the 5 -day BOD values will indicate the
proper quantity of seed. The table in
Figure 5 shows the 2/5 day ratio for each
of the seed concentrations used. When the
0.1% seed was used a low ratio was ob-
tained, indicating a lag period. When the
proper quantity of seed was used (0. 2 &
0. 5%) the ratio was higher and remained
essentially constant.
T 1"
5 DAY
BOD
2 DAY
5 DAY
1 1 I I 1 I 1 _-L.
I ? 1 4 «i (-780 10
DAYS INCUBATION @ ?0°C
EFFECT OF SEED CONCENTRATION ON THE B 0 D OF
GLUCOSE - GLUTAMIC ACID PRIMARY STANDARD
HGURF 5
8-10
-------
BOD: Effect of Variables
The actual value of the 2/5 day ratio de-
pends upon the k rate and is a. function of
the biological availability of the waste ma-
terial. It is therefore necessary to de-
termine the proper seed concentration for
the particular waste-seed combination to
be used.
As a rule of thumb for the seed concen-
tration, at least 0. 6 mg/1 of the 5 day BOD
should be due to the seed inoculum. Larger
seed demands are not objectionable but
tend to reduce the amount of oxygen avail-
able for the waste itself.
F Algae
When large numbers of algae are present
in stream waters, they produce significant
changes in the oxygen content. Under the
influence of sunlight oxygen is given off,
while during hours of darkness the algae
utilizes oxygen in their respiration. These
diurnal fluctuations in DO influence the
BOD of the stream water. In sewage sta-
bilization ponds the BOD of the waste is
satisfied primarily through the production
of DO by algae.
When stream samples containing algae are
incubated in the laboratory the algae sur-
vive for a time, then die because of the
lack of light. Short term BOD determi-
nations may show the influence of oxygen
production by the algae. When the algae
are dead they contribute to the total or-
ganic content of the sample and increase
the BOD. Therefore samples incubated
in the dark may not be representative of
the deoxygenation process in the stream,
since the benefits of photosynthesis are
lacking. Conversely, samples incubated
in the light, under conditions of continual
photosynthesis, will yield low BOD values.
The influence of algae in the BOD is one
of the most difficult variables to evaluate.
More research is needed to develop satis-
factory methods for the accurate deternii-
nation of BOD in the presence of large
numbers of algae.
IV TOXICITY
A Effect
Since satisfaction of the BOD is accom-
plished through the action of micro-
organisms, the presence of toxic sub-
stances will result in depression of the
oxidation rate. In many cases, toxicity
will produce a lag period, until resistant
organisms have evolved. Figure 6 shows
the effect of cyanide on the BOD curve. A
prominent lag period is exhibited in the
2 ppm curve, while at 10 ppm the lag ex-
tends beyond the fifth day.
Heavy metals reduce biological activity
and resistant organisms are seldom de-
veloped. The effect of copper and chromi-
um are illustrated in Figure 7.
012 34
Time in Days
Kffect of Cyanide on BOD of Domestic Sewage
(2% Sewage in Formula C Dilution Water)
-------
BOD: Effect of Variables
B Detection
In laboratory determinations of BOD the
absence of toxic substances must be es-
tablished before the results can be accepted
as valid. Comparison of BOD values for
ranging dilutions of the waste will indicate
the presence or absence of toxicity. In the
following table the calculated BOD for the
dilutions show higher values in the more
dilute concentrations. It is apparent that
toxicity is present and that it becomes
less with higher waste dilution, resulting
in more complete oxidation.
Table I
Dilution
10%
5%
2%
1%
0.5%
Depletion
3.51
4.53
2.80
1.52
0.74
5 day BOD
35
91
140
152
148
VII NITRIFICATION
A Mechanism
The oxidation process, as exemplified by
the equation:
y = L(l-10~kt)
involves the oxidation of carbonaceous
matter:
O
C H O
x y z
C0
H20
The constant k is normally high, giving
nearly complete oxidation in a few days.
When nitrogenous material is present its
oxidation can be shown as:
O2 O2
NH - N0 - N0
The rate constant is usually less than in
the case of the carbonaceous matter.
Under some circumstances these two oxi- "
dations can proceed simultaneously and
the resultant BOD curve will be a compo-
site of the two reactions. Normally, how-
ever, the nitrification stage will not begin
until the carbonaceous demand has been
partially satisfied, yielding a curve simi-
lar to that in Figure 8. Theriault suggested
that carbon dioxide, produced in the first
stage, is necessary for efficient oxidation
of the nitrogenous material. Mathemati-
cally the reactions can be described by the
equation:
y = L[ a (l-!0"kt) + b (l-lo"kt)]
where:
a = high rate constant for carbonaceous
b = low rate constant for nitrogenous
k1 = velocity constant for carbonaceous
k. = velocity constant for nitrogenous
Nitrification occurs most often in effluents
and streams which have undergone partial
oxidation of the waste components. In these
cases the total BOD (carbonaceous + nitro-
genous) is not representative of the degree
of treatment required.
Since nitrification represents a demand
on the oxygen resources of the receiving
stream, it should be recognized as part
T T
1 r
-i r
EFCECT OF N.TO FICA^'ON 0^ B 0 D
^'CURE 8
-------
BOD: Effect of Variables
of the total demand of the waste. Because
of the variable occurrence of the reaction,
however, comparison of BOD values is
usually restricted to the carbonaceous de-
mand.
B Detection and Measurement
When nitrification takes place during the
BOD incubation period, its presence can
generally be detected by the characteristic
shape of the BOD curve (Figure 8). A
significant upward swing of the curve after
initial rate has been established is an indi-
cation of nitrification.
,To determine the extent of nitrification it
/is necessary to measure the concentrations
/ of ammonia, nitrite nitrogen, and nitrate
/ nitrogen which corresponds to the BOD
t values obtained. Suitable correction for
i
\ these forms of nitrogen will yield the true
\ carbonaceous demand.
C Inhibition
The carbonaceous demand can be obtained
by preventing the nitrification process.
Because of the nature of the organisms
which are involved in the nitrogen reactions,
inhibition may be accomplished by pasteuri-
zation of the sample at 143op or acidifi-
cation to pH 2-3 with subsequent neutrali-
zation.
/•Ill EFFECT OF DILUTION
When a series of dilutions are made on a
BOD sample usually the results vary to the
extent that only an approximate BOD value
is obtained.
A For example, in Table 2, 1%, 2% and 4%
dilutions were made on a sample. The
4% dilution became anaerobic before the
end of 5 days. The 5-day BOD of the 1%
dilution was 270 and that of the 2% dilution
was 245.
B Statistically one value is more reliable
than the othe r.
Dilution
1%
2%
DO
5.5 mg/1
3.3 mg/1
2.2 mg/1
The difference of 1% dilution is 2. 2 mg/1.
Therefore, 2. 2 X 100 = 220 mg/1 BOD.
The calculated BOD of 220 mg/1 is closer
to 245 mg/1 than to 270 mg/1 BOD; there-
fore, the 245 mg/1 BOD is considered the
more valid BOD for this sample.
Table 2
INTERPRETATION OF BOD DATA
Sample
Initial
Final:
1% dilution
2% dilution
4% dilution
DO
8.2
5.5
3.3
0. 0
Depletion
-
2. 7
4. 9
BOD
-
270
245
Complete! - •
3-13
-------
USE OF BECKMAN CARBONACEOUS ANALYZER FOR
DETERMINING ORGANIC CARBON IN WATER
Robert T. Williams
I INTRODUCTION
A History of Carbon Analyses
In the wake of a rapid population growth,
and the increasing heavy use of our
natural waterways, the nation, and indeed
the TO rid, is presented with the acute
problem of increased pollutional loads on
streams, rivers and other receiving
bodies. This has resulted in a growing
awareness of the need to prevent the
pollution of streams, rivers, lakes and
even the oceans. Along with this aware-
ness has developed a desire for a more
rapid and precise method of detecting and
measuring pollution due to organic
materials.
B The Methods
In the past, two general approaches have
been used in evaluating the degree of
organic water pollution.
1 The determination of the amount of
oxygen or other oxidants required to
react with organic impurities.
2 The determination of the amount of
total carbon present in these impurities.
C Oxygen Demand Analyses
The first approach is represented by
conventional laboratory tests for determin-
ing Chemical Oxygen Demand (COD) and
Biochemical Oxygen Demand (BOD). One
of the principal disadvantages of these
tests is that they are limited primarily to
historical significance, that is, they tell
what a treatment plant had been doing,
since they require anywhere from two
hours to five days to complete. Since up
to now no faster method has been available,
traditional BOD and COD determinations
have become accepted standards of mea-
sure in water pollution control work even
though they are essentially ineffective for
process control purposes.
Until the introduction of the Carbonaceous
Analyzer, all methods taking the second
approach, the total carbon method of
evaluating water quality, also proved too
slow.
II THE ANALYSIS OF CARBON
A Pollution Indicator
Now the carbonaceous analyzer provides
a means to determine the total carbon
content of a dilute water sample in approx-
imately two minutes. With proper sample
preparation to remove inorganic carbonates,
the instrument determines the total organic
carbon content in the sample.
B Relationship of Carbon Analysis to BOD
and COD
This quantity varies with the structure
from 27 percent for oxalic acid through
40 percent for glucose to 75 percent for
methane. The ratio of COD to mg carbon
also varies widely from 0. 67 for oxalic
acid through 2. 67 for glucose to 5. 33 for
methane. Representative secondary sewage
effluents have given a ratio of COD to
carbon content of between 2. 5 and 3. 5
with the general average being 3.0.
The BOD, COD and carbon contents of
these and some other representative
compounds are summarized in the follow-
ing table.
*Leader, Analytical Services Group, Advanced Waste Treatment Research Program, Basic and
Applied Sciences Branch, DWS&PC, SEC. Reviewed December 1965.
CH. MET. 24.4. 65
9-1
-------
Use of Beckman Carbonaceous Analyzer
Sample
Stearic Acid - C 18H36O2
Glucose - CrH, 9OR
6126
Oxalic Acid - C2H2O4
Benzoic Acid - C~H,,O0
762
Phenol - CCH.O
6 6
Potassium Acid Phthalate
KHC H O
Salicylic Acid - C-HgO,,
Secondary Effluent, Clarified
n ii "
11 n 11
5 -Day
BOD-mg/mg
. 786
. 7 3
. 14
1.38
. 05 to 2. 1 de-
pending upon
concentration
.95
1. 25
13*
23*
4*
COD-
mg/mg
2.91
1.07
.18
1.97
2.36
1. 15
1. 60
67*
36*
% Carbon
76
40
27
69
77
47
61
21*
1 2-'-
7=:=
cln units of mg/1
III THE CARBON ANALYZER
A Principle of Operation
Basically the carbonaceous analyzer con-
sists of three sections - a sampling and
oxidizing system, a Beckman Model 315
Infrared Analyzer, and a strip-chart
recorder.
Carbona( coin Aniilvzer schematic
N oxygen carrier from cylinder LJcoohnp fjn
A micro sample (20 (j.1) of the water to be
analyzed is injected into a catalytic com-
bustion tube which is enclosed by an elect n<-
furnace thermostated at 950°C. The water
is vaporized and the carbonaceous material
is oxidized to carbon dioxide (CO^) and
steam in a carrier stream of pine oxyivn
The oxygen flow carries the steam and
CO2 out of the furnace where the steam ih
condensed and the condensate removed.
The CO2, oxygen and remaining water \apor
enter an infrared analyzer sensiti/.od to
provide a measure of CO2 The output of
the infrared analyzer is recorded on a strip
chart, after which, the curve produced can
be evaluated by comparing peak height with
a calibration curve based upon standard
solutions. Results are obtained directl\ in
milligrams of carbon per liter.
B Application
(1)
Results show that the method is applicable
for most, if not all, water-soluble organic
compounds -- including those that contain
sulfur, nitrogen, and volatiles.
Nonvolatile organic substances can be
differentiated from volatiles, such os
carbon dioxide or light hydrocarbons, by
9-2
-------
Use of Beckman Carbonaceous Analyzer
determination of carbon both before and
after the sample solution has been blown
with an inert gas. Investigators at the
Dow Chemical Company have reported' '
a method to be used for the determination
of the volatile constituents of a liquid
sample. The method involves the use of a
diffusion cell and sampling of both the
contained liquid and overhead vapor. The
combined values can then be related to
the carbonaceous material originally in
the liquid.
C Sample Preparation
The Carbonaceous Analyzer is often
referred to as a total carbon analyzer
because it provides a measure of all the
carbonaceous material in a sample, both
organic and inorganic. However, if a
measure of organic carbon alone is de-
sired, the inorganic carbon content of
the sample can be removed during sample
preparation.
1 Removal of inorganic carbon
The simplest procedure for removing
inorganic carbon from the sample is
one of acidifying and blowing. A few
drops of HC1 per 100 ml of sample will
normally reduce pH to 2 or less, re-
leasing all the inorganic carbon as CO2-
Five minutes of blowing with a gas free
of CC>2 sweeps out the CC>2 formed by
the inorganic carbon. Only the organic
carbon remains in the sample and may
be analyzed without the inorganic inter-
ference.
2 Volatile carbonaceous material
If volatile carbonaceous materials are
present in the sample, the procedure,
using a diffusion cell, must be followed
since light hydrocarbons would be lost
in the blowing process.
3 Dilute samples
If the sample is dilute (less than 100
mg/liter carbon) and is a true solution
(no suspended particles) no further
preparation is required.
4 Samples containing solids
If the sample contains solids and/or
fibers which are to be included in the
determination, these must be reduced
in size so that they will be able to pass
through the needle which has an opening
of 170 microns (needles having larger
openings may be obtained if necessary).
In most cases, mixing the sample in a
Waring Blender will reduce the particle
size sufficiently for sampling.
IV PROCEDURE FOR ANALYSIS
A Interferences
Water vapor resulting from vaporization of
the sample, causes a slight interference in
the method. Most of the water is trapped
out by the air condenser positioned immed-
iately after the combustion furnace. How-
ever, a portion of the water vapor passes
through the system into the infrared de-
tector and appears on the strip chart as
carbon. The water blank also appears on
the standard calibration curve, and is
therefore removed from the final calcu-
lation. In tests of solutions containing the
following anions: NO" Cl", SO"2, PO^3,
no interference was encountered with con-
centrations up to one percent.
B Precision and Accuracy
The recovery of carbon, from standard
solutions is__98T5_: 100. 5~percent^ The
minimum detectable concentration using
the prescribed operating instructions is 1
mg/1 carbon. Generally, the data are
reproducible to + 1 mg/1 with a standard
deviation of 0. 7 mg/1 at the 100 mg/1 level.
V APPLICATIONS
Several of the many research and industrial
applications of the Carbonaceous Analyzer
are listed below:
A Determine the efficiency of various waste -
water renovation processes, both in the
laboratory and in the field.
9-3
-------
Use of Beckman Carbonaceous Analyzer
B Compare a plant's waste outlet with its
water inlet to determine the degree of
contamination contributed.
C Monitoring a waste stream to check for
product loss.
D Follow the rate of utilization or organic
nutrients by micro-organisms.
E To detect organic impurities in inorganic
compounds.
VI ADVANTAGES OF CARBON ANALYZER
A Speed
The Carbonaceous Analyzer's most
important advantage is its speed of
analysis. One analysis can be performed
in 2-3 minutes. This speed of analysis
brings about another advantage, economy
of operation. Working with dilute samples,
one man can run ten to twenty carbon
determinations per hour. This is probably
more than the number of COD or BOD
tests that can even be started, much less
completed, in the same period of time.
B Total Carbon
Another advantage is that the measure of
carbon is a total one. The oxidizing
system of the analyzer brings about com-
plete oxidation of any form of carbon. No
compound has been found to which the
method is inapplicable.
VII CONCLUSIONS
The Carbonaceous Analyzer provides a
rapid and precise measurement of organic
carbon in both liquid and air samples. It
should be found useful for many research
and industrial applications, a few of which
have been mentioned.
Because of its rapidity it may be found more
useful than the more time-consuming BOD
and COD measurements for monitoring
industrial waste streams or waste treatment
processes.
REFERENCES
1 Van Hall, C. E. , Safranko, John and
Stenger, V. A. Anal. Chem. 35,
315-9. 1963.
2 Van Hall, C. E., and Stenger, V. A.
Draft of Final Report - Phase I - Con-
tract PH 86-63-94, Analytical Research
Toward Application of the Dow Total
Carbon Determination Apparatus to the
Measurement of Water Pollution.
3 Van Hall, C. E. , Stenger, V. A.
Beckman Reprint - R6215. Taken from
Paper Presented at the Symposium on
Water Renovation, Sponsored by the
Division of Water and Waste Chemistry.
ACS in Cincinnati. Jan. 14-16, 1963.
9-4
-------
NUTRIENTS IN WATER - THE PROBLEM
Michael E. Bender*
I INTRODUCTION
A Nutrients of importance include macro-
nutrients: those needed in large quantities,
and micronutrients: those needed in
small amounts.
B These nutrients are important because
they promote biological responses which
interfere with some desired use of the
water by man.
C Other factors (e.g. temperature, light)
affect the use of these nutrients and
should be considered in an evaluation of the
effects of nutrients upon the aquatic
environment.
B Shelford's "law" of tolerance: survival
of an organism can be controlled by the
quantitative or qualitative deficiency or
excess with respect to any one of several
factors which may approach the limits of
tolerance for that organism.
C Qio "law": with a temperature increase
of 10 degrees centigrade metabolic pro-
cesses (rates) are approximately doubled.
IV The process of photosynthesis is the
fixation of the sun's energy with the pro-
duction of organic matter by plants.
A The general reaction is given below:
II Algae, bacteria, and aquatic plants are
the forms of life v/hich nutrients affect most
directly.
A Algae are of Several Types
1 ^Phytoplankton are small algae suspend-
gd in the water and form the basis of
productivity in the aquatic environment.
2 Benthic algae are those forms anchored
to substrates of rock and bottom
materials.
B Aquatic plants are of several types. In
general they may be referred to as rooted
or floating forms.
-ES-SRend tQ organic nutrieais_intr.odu.ae,cL
into w.ater. A_u|Qtrnphic ba.cteria may re-
spond and grow due to inorganic nutrient
sources.
Ill BIOLOGICAL LAWS
A Liebig's "law" of the minimum: the
essential material available in amounts
most closely approaching the critical
minimum needed will tend to be the limiting
C0
B Chlorophyll contains basically C, O, H,
N and Mg, and in general makes up
about 5% of the dry weight of algal cells.
V MEASUREMENT OF PHOTOSYNTHESIS
A Oxygen production can be used as a
measure of photosynthesis because for
each mole of CO2 reduced to organic
carbon one mole of free oxygen is liberated.
1 The value of O2/CO2 has been found
experimentally to be 1.25 rather than
1.0.
B CO2 Assimilation
1 The CO2 taken up by algae does not all
originate from the dissolved gas. Some
algae can use bicarbonate directly as
a source of carbon.
2 Hence measurement of CO? uptake from
water is a complicated problem which
must consider pH,
centrations.
HCO
3'
and CO* con-
*Former Biologist, DWS&PC Training Activities,
W.RE.ntr. 2. 12.63
SEC. Reviewed December 1965.
10-1
-------
Nutrients in Water - The Problem
C Fixation of Carbon-14
1 The use of C*4 as a tracer of C
12
in
plant metabolism and productivity
estimation has been widely used since
the early nineteen fifties.
14
In this method a known amount of C
is added to the water and after a period
of time the proportion of C14 in the
plant cells to C14 added is found. The
amount of carbon assimilated is then
estimated from the following equation.
activity of
phytoplankton
activity of
C14O* added
(K) =
total carbon
assimilated
total carbon
available
Where K is a constant relating to the
slower uptake of C^4.
The total carbon available is deter-
mined chemically.
D Uptake of Mineral Nutrients
1 The measurement of depletion of
nutrients in solution has been tried
but found unreliable.
E Chlorophyll
1 The quantity of chlorophyll present
has been found to bear some relation
to productivity but not a reliable one.
VI Nutrients of signifance in the growth and
production of algae and plants are discussed
below.
A Carbon
1 Sources
a Gaseous CO_
co
2 Effects of the removal of carbon upon
the water
a Lowered pH
b Deposition of CaCO
-------
Nutrients in Water - The Problem
be the limiting growth factor in these
populations.
Inorganic micronutrients - Many elements
are needed in very small quantities by
algal cells. Some of these have a known
function in algal metabolism; others do
not.
1 Mg is a cation of major importance in
the chlorophyll molecule.
These interferences include taste and odor,
filter clogging, and oxygen depletion which
may result in fish kills and subsequent
nuisance conditions.
B Benefits derived from productivity include
oxygenation and hence self purification in
polluted areas. Increased biological
populations result from increased pro-
ductivity and this may lead to increased
numbers of desirable fish species.
2 Co is known to be necessary for vitamin yjjj CYCLE OF NUTRIENTS
B
12'
3 Mn is necessary for several enzyme
systems.
4 Mo, V, Zn, and Cu are necessary but
these functions are not as well known.
A Once nutrients enter a body of water they
are cycled through a food chain.
B Factors affecting this food chain (e.g.
toxicity, removal) will affect the concentra-
tion and distribution of the nutrients.
F Organic Micronutrients
1 Some 40% of algae investigated have
been shown to require vitamins. The
following breakdown of vitamins needed
has been established for these algae:
a B12 80%
b Thiamine 53%
c Biotin 10%
2 Algae can use many organic compounds
/ as sources of N, C and P, The im-
;v portance of these compounds in natural
\ waters seems small.
VII PROBLEMS AND BENEFITS DERIVED
FROM ALGAL PRODUCTION
A The major problems occur from inter-
ference with a desired use for water.
REFERENCES
1 Lewin, Ralph A. Physiology and Bio-
chemistry of Algae. Academic Press.
1962.
2 Odum, Eugene P. Fundamentals of
Ecology. W. B. Saunders Co. 1959.
3 Odum, H. T. Primary Production in
Flowing Waters, Limnology and
Oceanography. 1(2): 102-117. April
1956.
4 Ryther, John H. The Measurement of
Primary Production. Limnology and
Oceanography, 1(2): 72-84. April
1956.
5 Verduin, Jacob. Primary Production in
Lakes. Limnology and Oceanography.
1(2): 85-91. April 1956.
10-3
-------
SOURCES AND ANALYSIS OF ORGANIC NITROGEN
F. J. Ludzack*
I INTRODUCTION
A Organic nitrogen refers to the nitrogen in
combination with any organic radical.
For sanitary and civil engineering the
main interest is the nitrogen contained in
proteins, peptides, amines, amino acids,
amides and other protein compounds of
animal or vegetable origin. Analytical
methods are designed to estimate these
and may not include certain other forms
of organic nitrogen such as nitro or nitrile
nitrogen.
B Most nitrogen compounds are characterized
by rapid conversion from one form to
another by biological and chemical action.
Hydrolysis, deamination, peptide forma-
tion, and other reactions may appreciably
alter the original form of sample nitrogen
within a short time.
II SOURCES OF ORGANIC NITROGEN
A Natural Origin
1 Dead animal and plant residues
2 Animal wastes - urea, feces
3 Autotrophic organisms - algae, s.bact.
4 Heterotrophic organisms ~f*—*•-*-*•
B Industrial Origin
1 Food processing wastewater-meat,
milk, vegetables.
2 Pharmaceutical wastes, antibiologicals.
3 Plastics - polyamides, nitriles.
4 Chemical intermediates or products.
5 Dye industry - azo, nitro.
Ill ANALYSIS OF ORGANIC NITROGEN
A No general procedure is likely to give high
analytical recovery on all forms of organic
nitrogen. Time, temperature of digestion,
catalyst and technique may require adjust-
ment for optimum performance.
B The organic nitrogen determination was
designed for items listed in I. A. and their
breakdown products. Other forms of
organic nitrogen may not give good yields
by the same technique.
C Nature and composition of extraneous ma-
terials affect analytical recovery. High
salt concentrations may raise digestion
temperature. Fats and carbohydrates may
use up acid during oxidation resulting in
insufficient acid for nitrogen digestion.
D Analytical recovery should be checked for
unusual samples to evaluate suitability of
the test routine.
E McKenzie and Wallace observations (Aust.
Jour, of Chem. 7. No. 1. 55-70, 1954).
1 Digestion temperature is critical. From
380 to 390°C usuaUy gives high analytical
recovery on the more refractory nitrogen
compounds of natural origin. Nitrogen
losses occur above 420°C. The tem-
perature creeps upward during digestion.
2 The optimum temperature is associated
with a digestion mix containing 1 g of
potassium sulfate for each ml of sul-
furic acid.
.^^^Mercury is the best catalyst but the
-------
Sources and Analysis of Organic Nitrogen
4 Tellurium almost as good as Hg, no
sulfide pptd. required.
5 H2C>2 is the only known oxidant besides
hot 62804 that does not tend to oxidize
organic nitrogen to N2. Its use is not
recommended unless Hg cannot be used.
It is tedious, time consuming, and tends
toward low yields.
6 About 7.3 g of H2SO4 was needed to
oxidize 1 g of carbohydrate. About 9 g
for each g of fat. Acid addition should
allow for acid use by extraneous
compounds.
F Standard Methods, APHA llth Ed.
The procedure basically is that of Morgan,
Lackey, andGilcreas(Anal. Chem. 11, 833,
1957). This modification was based on "
the method of McKenzie and Wallace.
\
1 The digestion mix has 0.67 g of
potassium sulfate per ml 112804 cor- j
responding to a digestion temperature j
of about 360°. '
2 The digestion mix tends to ppt. on
storage. It must be diluted to hold
the sulfate in solution at room tempera-
ture.
3 The procedure appears effective for
general use but the digestion tempera-
ture is low for refractory compounds
unless the digestion time is lenghtened
which raises temperature.
11-2
-------
DETERMINATION OF PHOSPHATES IN WATER
J. M. Cohen*
I INTRODUCTION
A Why concern about phosphates in water?
1 Nutriait for algae which can cause taste
and odors ancTcan inteffere"with sand
filtration.
2 Polyphosphates can interfere with
chemical coagulation of water.
B Source of Phosphates in Water
1 Geological
2 Agricultural fertilizers
3 Sewage
a human wastes
b synthetic detergents
c biological protoplasm
4 Industrial wastes
5 Additive for corrosion control
II CHEMISTRY OF POLYPHOSPHATES
To the nonspecialist the chemistry of the
phosphorus compounds is largely an unknown
area. Increased commercial use of polyphos-
phates makes it important that chemists
know more of the chemistry of these materials.
A Most common forms of phosphorus are
. the orthophosphates derived from substi-
/ tution of the replaceable hydrogens in
phosphoric acid (Figure 1).
B When orthophosphates or mixtures of
orthophosphates are heated, molecular
water is withdrawn and a new class of
phosphates are formed, called dehydrated or
condensed phosphates. These compounds
belong to the more general class of
phosphates called polyphosphates.
Monos odium
Dihydrogen
Orthophosphate
Na3P°4
Trisodium
Ortho-
phosphate
Na2HP04
Dis odium
Monohydrogen
Orthophosphate
Nomenclature for Orthophosphates
Figure 1
1 A chemical definition of a crystalline
polyphosphate is those phosphates in
which two phosphorus atoms are linked
through an oxygen atom as shown in
Figure 2.
o~~ o o
I I
o=p—o— o—P—o—P—o
o— o— o —
Orthophosphate Pyrophosphate
o— o o—
II
O=P— O—P—O—P=O
o— o— o —
Triphosphate
Schematic Formulae of Phosphates
Figure 2
^Chemical, Basic and Applied Sciences Branch, DWSPC, SEC. Reviewed December 1965.
CH. PHOS. 2a. 12.63 12-1
-------
Determination of Phosphates in Water
2 Polyphosphates are formed by abstract-
ing molecular water by heating mono
and dibasic orthophosphates as shown
in Figure 3.
fv\
• 3) Single largest consumer of polyphos-
'-J phates are synthetic detergents which
may contain 40 to 60% of a polyphos-
phate. " ~
',,a Practically all of the annual pro-
i, duction of 650, 000 tons is eventu-
ally discharged to a natural water.
C Polyphosphates have the property of rehy-
drating or reverting to the orthophosphate
forms.
1 The rate at which such reversion oc-
curs is important, since the character-
istic behavior of polyphosphates is lost
as rapidly as it reverts to the ortho
form.
2 The rate of reversion of tripolyphos-
phate in Ohio River water is seen to be
very slow, as shown in Figure 4.
3 Other factors, as shown in Figure 5,
greatly influence the reversion rate.
n(NaH,PO,) A2°° " 40°°,C (NaPO J + nH?O
£t 4 o n ^
Monos odium
Orthophosphate
2Na2HP04 - NaH2P04-
S odium
Trimetaphosphate
Disodium Monosodium
Ortho- Ortho-
phosphate phosphate
2H2°
Pentas odium
Tri-
phosphate
Preparation of a Polyphosphate
Figure 3
in OCCURRENCE OF PHOSPHATES IN WATE*R
A Types encountered as water contaminant
1 Orthophosphate
Source can be fertilizer, endproductof
polyphosphate reversion, an industrial
waste and geological.
2 Polyphosphate
Source can be synthetic detergents,
water conditioning and, to a lesser ex-
tent, certain biological compounds.
3 Biological Protoplasm
Phosphorus which has been incorpo-
rated in organisms during metabolism.
4 Organic Phosphate
Small amounts originate from human
wastes, from decomposed biological
fauna and industrial wastes.
B Types of Phosphate Analysis
The above types of phosphates can be de-
termined in a water sample in the follow-
ing manner.
1 Total phosphates
This analysis will yield all forms of
phosphate and will include:
a ortho
b poly
c biological
d organic
2 Total soluble phosphate
Analysis performed on filtered sample
will include:
a soluble ortho
b soluble poly
12-2
-------
Determination of Phosphates in Water
10
Figure 4
20
40
60 ' ' 180
Elapsed Time - days
200
220
240'
TRIPOLYPHOSPHATE REVERSION IN OHIO R. WATER
Factor
Approximate Effect on Rate
Temperature
PH
Enzymes
Colloidal Gels
Complexing Cations
Concentration
Ionic Environment in
the Solution
105 - 106 Faster from Freezing to Boiling
103 - 104 Slower from Strong Acid to Base
As much as 105 - 10B Faster
As much as 104 - 105 Faster
Several Times Faster
Roughly Proportional
Severalfold Change
Major Factors Affecting the Rate at Which
Chain and Ring Phosphates Undergo Hydrolytic Degradation
Figure 5
-------
Determination of Phosphates in Water
c soluble organic
3 Ortho phosphate
This is the most common determination
and will theoretically determine only
this form to the exclusion of all others.
IV ANALYTICAL PROCEDURES
A Procedures most applicable to water
analysis are colorimetric methods whose
basic reactions are as follows:
1 Formation of phosphomolybdic com-
pound by adding ammonium molybdate
to form a yellow colored heteropoly
acid.
2 Reduction of the heteropoly acid with
some reductant, for example, stannous
chloride.
3 Measurement of the intensity of the
blue color and comparing with a standard
curve.
B Orthophosphates are determined by apply-
ing the above procedure.
C Polyphosphates are determined by analyz-
ing for orthophosphate before and after
acid hydrolysis:
ortho (after hydrolysis) - ortho (before hy-
drolysis) = Polyphosphate (as
4
D Total phosphates are determined by wet-
digesting the sample to convert all forms
to the ortho for which an analysis is then
made.
V VARIABLES IN ORTHOPHOSPHATE
ANALYSIS
A Ratio of acid to molybdate must be main-
tained constant at the optimum.
B Amount of molybdate reagent is not criti-
cal, so long as the final acidity is about
0. 2 - 0. 4 N sulfuric acid.
C Amount of reductant is also not critical,
although too much reagent can lead to tur-
bidity on standing.
D Time for color development must be
closely controlled for both the standard
and sample.
E Temperature produces about a 1% in-
crease in color for each 1°C rise.
F Interferences
1 These ions must be absent - silver,
arsenate, barium, dichromate, mer-
curic, mercurous, molybdate, lead,
chlorostannate, zirconium, tungstate,
silicate and nitrite.
2 Other ions interfere but can be present
in varying concentrations before intro-
ducing an error greater than 2%.
VI RECENT MODIFICATION IN ANALYTICAL
PROCEDURE^)
A Modification avoids these two weaknesses
of the above method.
1 Partial hydrolysis of the polyphosphates
because of the acid conditions of the
test.
2 Interfering substances in natural waters.
3 Turbidity must be removed.
B Modification consists of extracting the re-
action product of phosphate and molybdate
with a 1 + 1 mixture of benzene and iso-
butanol. Reduction of the heteropoly acid
is performed in the organic layer.
VII RESULTS AND SENSITIVITY
A In water analysis results are generally
expressed as PO4 although other expres-
sions, such as P or ?2O5, may be used.
12-4
-------
Determination of Phosphates in Water
B Colorimetric methods are highly sensitive
and will detect down to 0. 002 ppm.
C In the range of 0.2 - 1.0 ppm a precision
of + 10% can be obtained.
REFERENCES
1 Moss, H. V. (Chairman, AASGP Com-
mittee) "Determination of Orthophos-
phate, Hydrolyzable Phosphate and
Total Phosphate in Surface Waters".
JAWWA, _50:1563. December 1958.
2 Quimby, Oscar T. "The Chemistry of
Sodium Phosphate". Chemical Reviews,
50:141. February 1947.
3 Standard Methods for the Examination of
Water and Wastewater, llth Edition,
APHA, AWWA, WPCF. 1961.
12-5
-------
DETERMINATION OF ORTHO AND POLYPHOSPHATE BY
MOLYBDENUM BLUE-STANNOUS CHLORIDE METHOD
R. J. Lishka*
I COLLECTION OF SAMPLE
Collect sample in the prescribed manner and
analyze as soon as possible to minimize con-
version of polyphosphate to orthophosphate.
II APPARATUS
Spectrophotometer or filter photometer,
use at 690 millimicrons.
for
Add 155 ml of cone. H2SO4 slowly to
400 ml of distilled water and cool.
Add the molybdate solution to the sulfuric
acid solution (never in reverse) and
dilute to 1.0 liter.
E Stannous chloride solution - Dissolve 2.5
gms of fresh SnClg' 2H2O in 100 ml of
reagent grade glycerine. Heat in water
bath and stir to dissolve.
Ill REAGENTS
A Phenolphthalein indicator - Dissolve 2.5
gms of phenolphthalein powder in 250 ml
of ethyl alcohol, add 250 ml of distilled
water, then add 0. 020N NaOH dropwise
to a faint pink color.
B Sulfuric acid -Add 310 ml of cone. H2SO4
slowly to about 600 ml of distilled water.
Cool to room temperature and dilute to
1.0 liter.
IV PROCEDURE
Preliminary Cleaning of Glassware. Because
laboratory detergents contain phosphate which
cannot be easily rinsed off, a preliminary
treatment with phosphate reagents is necessary
to remove that adsorbed on the glass sur-
faces. After the glassware has been properly
cleaned it should be reserved for this test only.
A Clean and thoroughly rinse all glassware
in the usual manner.
C Standard Phosphate
1 Stock standard phosphate solution
(0. 50 mg PO4 per ml). Dry a portion
of reagent grade potassium dihydrogen
phosphate overnight at 103°C before
use. Dissolve 0.7164 gms of the
KH2PO4 in distilled water and make up
to 1.0 liter.
2 Working standard phosphate solution
(0.005mg/PO4 per ml). Dilute 10.0
ml of the stock standard phosphate to
1. 0 liter with distilled water. Protect
this solution from the light and make
up fresh each month.
B Before use, treat each container which
will come in contact with the sample as
follows:
1 Fill each container with distilled water
and add 2. 0 ml of ammonium molybdate
solution, reagent III. D, and mix.
2 Add 1 ml of sulfuric acid solution,
reagent III. B, and 1 ml of Stannous
chloride solution, reagent III. E. Mix
thoroughly and allow to stand for 10-
15 minutes. Make certain that all
internal surfaces of the containing
vessels come in contact with the reaction
mixture. Discard the reaction mixture
and rinse each container with distilled
water.
D Ammonium molybdate solution - Dissolve
25.0 gms of (NH4)6Mo7O24-4H2O in 175
ml of distilled water.
Once this cleaning procedure has been per-
formed the glassware can be reused without
repeating the treatment unless it has been
cleaned with detergent.
*Chemist, Analytical Reference Service, Training Program, SEC. Reviewed December 1965.
CH. PHOS.lab. 1. 12. 63 r>-(j
-------
Determination of Phosphate (Ortho and Poly)
Preparation of Standards (Photometric
Procedure) Observe room temperature.
Standards and samples must be run at
same temperature,
results.
+ 2 C, for reproducible
Add increasing volumes of working
standard phosphate reagent III. C. 2 to
several vessels (previously cleaned
according to IV. B). Add 1. 0 ml of
sulfuric acid, reagent III. B. Make up
to 50. 0 ml with distilled water.
The following table is suggested as
a convenient series for use with a
photometer.
ml of soln
III. C.2
0 (Blank)
0.5
1.0
2.0
5.0
10.0
mg PO4/50 ml
0
.0025
.0050
.0100
.0250
.0500
P04, mg/1
0 (Blank)
.05
0. 10
0.20
0.50
1.00
D Raw Water Sample (Photometric
Procedure)
1 Filter at least 200 ml of the sample to
remove turbidity.
2 Place 100 ml of the filtered sample
containing not more than 0. 050 mg PC>4
or an aliquot diluted to 100 ml in a
250 ml Erlenmeyer flask. Place several
glass beads or boiling stones in the flask.
3 Omit this step if the pH is below 8.2.
Otherwise
a add two drops of phenolphthalein,
reagent III. A
b add sulfuric acid solution, reagent
III. B, dropwise, to discharge the
pink color.
4 Add exactly 2.0 ml of sulfuric acid,
reagent III. B, and boil gently for one-
half hour. Add a small amount of
distilled water if volume drops below
25 ml during the boiling period.
5 Cool, make up to 100 ml with distilled
water and adjust to temperature of
standard curve. Place 50 ml in a
vessel (previously cleaned according to
step IV. B) and label Total Phosphate.
6 Place 25 ml, or an aliquot diluted to
25 ml, of the filtered sample from step
IV. D. 1 in a vessel (previously cleaned
according to step IV. B) and add 1. 0 ml
sulfuric acid, reagent III. B. Make up
to 50.0 ml with distilled water and label
Orthophosphate.
7 Add 2.0 ml of ammonium molybdate,
reagent III. D, to each sample and
standard. Mix thoroughly.
8 Add 5 drops (0.25ml) of stannous chloride
solution, reagent III. E. Mix well and
allow to stand for 20 (+5) minutes.
9 Read in spectrophotometer at 690 m|J..
Use the 0 standard to zero the instrument
10 Prepare calibration curve.
E Calculation
mgPO /I = mg P°4 X
1000
ml sample
mg polyphosphate/1 » mg total phosphate/I -
mg Orthophosphate/I
V NOTES
A The concentration of acid in the sample and
standards has a direct effect on the amount
of color produced. It is, therefore, im-
portant that addition of reagent III. B be
performed accurately.
B The phosphomolybdate blue produced by
^ the reaction is not stable and tends to
fade gradually after about 20 minutes. Most
accurate results are obtained when time
requirements are observed.
12-7
-------
Determination of Phosphate (Ortho and Poly)
C Temperature is another variable which
affects the color formation. Samples and
standards may be adjusted to room tem-
perature after boiling by allowing to
stand for several hours before addition of
reagents III. D and III. E.
D If the phosphate concentration in the
sample is completely unknown, several
dilutions of the sample may be run simul-
taneously and that aliquot used for final
reading which falls near the middle of the
standard series.
E Scrupulous care should be observed in
cleaning of the glassware prior to use. If
color appears in the blank, or a particular
standard appears out of line, phosphate
contamination is probably responsible.
The working standard phosphate solution,
reagent III. C. 2, should be kept in the
dark to prevent reduction of phosphate
concentration through algal growth.
In preparing reagent III. D, the molybdate
must be added to the sulfuric acid, never
the reverse. Reversing the addition
precipitates some of the molybdate, which
is then very difficult to dissolve.
12-f
-------
AMMONIA, NITRITES AND NITRATES
Betty Ann Punghorst*
I SOURCES AND SIGNIFICANCE OF
AMMONIA, NITRITES AND NITRATES
IN WATER
The natural occurrence of nitrogen compounds
is best demonstrated by the nitrogen cycle
( Figure 1) .
A Ammonia
1 Occurrence
Ammonia is a product of the micro-
biological decay of animal and plant
protein. In turn it can be used directly
to produce plant protein. Many fertili-
zers contain ammonia.
2 Significance
The presence of ammonia nitrogen in
raw surface waters might indicate do-
mestic pollution. Its presence in waters
used for drinking purposes may require
the addition of large amounts of chlorine
in order to produce a free chlorine
residual. The chlorine will first react , •
with ammonia to form chloramines be- \t
___for£ it exe*rts"Tts full pacferojidjij..effects i ''
(free chlorine residual). '
B Nitrite
1 Occurrence
Nitrite nitrogen occurs in water as an
intermediate stage in the biological de-
composition of organic nitrogen. Nitrite
formers ( nitrosomonas) convert ammonia
under aerobic conditions to nitrites. The
bacterial reduction of nitrates can also
produce nitrites under anaerobic condi-
tions. Nitrite is used as a corrosion
of larg e quantities indie at e s a source
~~
2 Significance
Nitrites are usually not found in_ surface
water to a^fe'aT'exfe'nT. The presence
C Nitrate
1 Occurrence
Nitrate formers convert nitrites under
aerobic conditions to nitrates (nitro-
bacter). During electrical storms,
large amounts of nitrogen ( N%) are
oxidized to form nitrates. Finally,
nitrates can be found in fertilizers.
2 Significance
Nitrates in waters usually indicate the
final stages of biological stablization.
Nitrate rich effluents discharging into
receiving_waters ?an^_under_j)roper en-
vironment conditions cause stressj^p
stream quality by groducingjilgal
blooms. Drinking waters containing
excessive amounts of nitrates can
causeinfant niethemoglobinemia.
II ANALYSIS OF AMMONIA, NITRITES,
AND NITRATES IN WATER
A Sample Collection
If a sample cannot be analyzed promptly,
several procedures may be followed in
order to preserve the sample.
1 Freezing will retard biological activity.
2 The addition of 1 ml of chloroform/100
ml sample should retard biological
activity.
3 The addition of 0. 8 ml cone H2SO4/1
liter of sample will also maintain the
nitrogen balance. However, it is
essential that the sample be neutralized
before proceeding with the analysis.
*Chemist, DWS&PC Training Activities, SEC. Reviewed December 1965.
CH.N.6. 11.64
13-1
-------
Ammonia, Nitrites and Nitrates
rterial Oxidatign)
Figure 1
The Nitrogen Cycle
13-2
-------
Ammonia, Nitrites and Nitrates
B Determination of Ammonia
1 Nesslerization
a Reaction
Nessler's reagent, a strong alkaline
solution of potassium mercuric iodide,
combines with NHg in alkaline solu-
tion to form a yellowish brown
colloidal dispersion.
3KOH
The intensity of the color follows the
Beer-Lambert Law and exhibits
maximum absorption at 425 m(i.
b Interferences
1 Nessler's reagent forms a pre-
cipitate with some ions (e.g.,
Ca++, Mg++, Fe+++, and S=).
These ions can be eliminated in
a pretreatment flocculation step
with zinc sulfate and alkali. Also
EDTA or Rochelle salt solution
prevents precipitation with Ca^+
or Mg++.
2 Residual chlorine indicates that
ammonia may be present in the
form of chloramines. The ad-
dition of sodium thiosulfate will
convert these chloramines to
ammonia.
3 Certain organics may produce an
off color with Nessler's reagent.
If these compounds are not steam
distillable, the interference may
be eliminated in the distillation
method.
4 If the turbidity and natural color
of the sample cannot be eliminated
with flocculation, it is then neces-
sary to use the following distilla-
tion method.
O + 7KI + 2H.O
It
Yellow-Brown C.~~
2 Distillation
a Reaction
1) The sample is distilled in the
presence of a phosphate buffer
at pH 7.2 - 7.4.
NH
+ H
H -t- NaHPO.
Buffer
NaH0PO.
Li Q
Na
pHmaintained be-
tween 7.2 - 7. 4
2) The ammonia in the distillate is
then measured by either of two
techniques.
a) Nesslerization is used for
samples containing less than
1 mg/ 1 of ammonia nitrogen.
b) Absorption of NHq by boric
acid and back titration with a
standard strong acid, is more
suitable for samples contain-
ing greater than 1 mg/1 of
NH N.
iJ
13-3
-------
Ammonia, Nitrites and Nitrates
NH + HBO -
..
3 Precision
On a synthetic sample containing 0.25
mg/1 of nitrite nitrogen, the Analytical
Reference Service (Water Minerals
Study, 1961) reported 125 results with
a standard deviation of + 0. 029 mg/1.
D Determination of Nitrate
1 Phenoldisulfonic acid
a Reaction
1) Phenoldisulfonic acid reacts with
nitrate to produce a nitro
derivative.
HSO,
OH
HSO
+ H + NO
13-4
-------
Ammonia. Nitrites and Nitrates
In alkaline solution the nitro derivative
rearranges to form a yellow-colored
compound which exhibits maximum ab-
sorption at 410 m|A.
OH
HSO
NO
KSOr
+ 3KOH-
S03H
COLORLESS
b Interferences
O
=N OK
S03K
YELLOW
1) Chloride ion under the acid con-
ditions of the test introduces a
negative interference.
6 Cl + 2NO0 +
2NOt+ 4H0O
£
Silver sulfate can be used to pre-
cipitate Cl" but due to incomplete
precipitation of Ag+, an off color
or turbidity is produced when the
final color is developed.
2) Nitrites in concentrations greater
than 0.2 mg/1 N introduce posi-
tive interference. However, in
most waters the concentration of
nitrite is insignificant as compared
to nitrate.
3) Color and turbidity may be re-
moved by using Al (OH) 3 suspen-
sion or by flocculation with ZnSO.
and alkali.
c Precision vx-t, „* & ^-^ '
On a synthetic sample containing 1. 1
mg/1 of nitrate N, the ARS Water
Minerals Study (1961) reported 118
results with a standard deviation of
+ 1. 119 mg/1.
2 Brucine
a Reaction
Brucine, a strychnine compound,
reacts with nitrate to form a yellow
compound which exhibits maximum
absorption at 410 mji. The reaction
according to the procedure as out-
lined in Standard Methods (p. 178)
does not follow Beer's Law. How-
ever, a recent modification by
Jenkins et al. (see reference 2) has
been developed. Conditions are con-
trolled in the reaction so that Beer's
Law is followed and concentrations
below 1 mg/1 nitrate nitrogen can
be determined.
b Interferences
1) Nitrite may react the same as
nitrate but can be eliminated by
the addition of sulfanilic acid to
the brucine reagent.
2) Organic nitrogen compounds may
hydrolyze and give positive inter-
ference at low (less than 1 mg/1)
nitrate nitrogen concentrations.
3) Residual chlorine may be elimi-
nated by the addition of sodium
arsenite.
c Precision
u
1) On a synthetic sample containing
1. 1 mg/1 of nitrate N, the ARS
Water Minerals Study (1961) re-
ported 21 results with a standard
deviation of + 0. 490 mg/1.
2) Jenkins, et al. report a standard
deviation of + 0. 0048 mg/1 on a
sample with a mean nitrate
nitrogen concentration of 0.287
mg/1.
3 Hydrazine reduction /,
, V-
'~ *V 2
A method using hydrazine to reduce
nitrate to nitrite followed by subsequent
13-5
-------
Ammonia, Nitrites and Nitrates
measurement of nitrite by diazotization
was recently reported by Fishman, et al.
(see reference 1). The procedure has
been successfully adapted to the Auto
Analyzer where a high degree of control
of reaction conditions can be achieved,
(see reference 3).
REFERENCES
1 Fishman, Marvin J., Skougstad, Marvin
W., and Scarbio, George Jr. Diazoti-
zation Method for Nitrate and Nitrite.
JAWWA. 56:633-638. May 1960.
2 Jenkins, David and Medsker, Lloyd L.
Brucine Method for Determination of
Nitrate in Ocean, Estuarine, and Fresh
Waters. Anal. Chem. 36:610-612.
March 1964.
3 Kamphake, L. J. Chemist, Engineering
Section, Basic and Applied Sciences
Branch, DWS&PC, Robert A. Taft
Sanitary Engineering Center. Personal
Communication.
4 Sawyer, Clair N. Chemistry for Sanitary
Engineers. McGraw-Hill Book Company,
Inc. New York. 1960.
5 Standard Methods for the Examination of
Water and Wastewater. APHA, AWWA,
WPCF. 1960.
Vs
-J
13-6
-------
AMMONIA DETERMINATION BY DIRECT NESSLERIZATION
R. J. Lishka*
A REAGENTS
1 Nessler reagent:
Dissolve 100 g mercuric iodide and
\j 70 g. potassium iodide in a small
' quantity of ammonia-free water and
I add to a cool solution of 160 g. sodium
\hydroxide in 500 ml ammonia-free
water. Dilute to 1 liter with ammonia-
free water.
2 Stock ammonium chloride solution:
Dissolve 3. 819 g anhydrous ammonium
chloride in ammonia-free water and
dilute to 1 liter.
1. 00 ml = 1. 00 mg N
3 Standard ammonium chloride solution:
Dilute 10 ml of stock ammonium
chloride solution to 1 liter with
ammonia-free water.
1. 00 ml = 0.0100 mg N
4 Zinc sulfate solution:
Dissolve 100 g ZnSO4 • 7 H2O in
ammonia-free water and dilute to 1
liter.
5 Sodium hydroxide solution:
Dissolve 240 g NaOH in 500 ml of
ammonia-free water and dilute to 1
liter.
6 Rochelle salt solution:
Dissolve 50 g K NaC4H4Og- 4 H2O
in 100 ml ammonia-free water. Boil
off 30 ml of the solution-'to remove
any ammonia present in the salt. After
cooling, dilute to 100 ml.
B PROCEDURE
Add 1 ml zinc sulfate solution to 100
ml of sample; mix thoroughly; add
0. 5 ml hydroxide solution; again mix
thoroughly.
After precipitate has settled, filter
the sample through Whatman No. 1
paper and discard the first 25 ml of
the filtrate.
Take 50. 0 ml, or an aliquot diluted to
50. 0 ml with ammonia-free water,
add 2 drops of Rochelle salt solution,
and mix well.
Prepare a series of standards con-
taining 0. 0, 0.5, 1.0, and 3. Oml of
standard ammonium chloride solution.
Dilute each to 50 ml with ammonia -
free water.
Add 1 ml Nessler reagent to each
standard and sample and mix well.
After 10 minutes, measure the ab-
sorbance in a spectrophotometer at
425 m|jL, using 1 cm. cells. Use the
0. 0 standard as a reference.
Plot the standard curve and determine
the mg of N in the sample.
C CALCULATION:
mg ammonia N X 1000
mg/1 ammonia N = — l sample nesslerized
*Chemist, Analytical Reference Service, Training Program, SEC. Reviewed December 1965.
CH.N.Iab. 1. 12. 63 13-7
-------
Ammonia Determination by Direct Nesslerization
AMMONIA DETERMINATION DATA SHEET
Absorbance ^TTT ,
mgNH_/N
reading 6 3
0.5 ml standard soln. 0.005
1.0 ml standard soln. 0.010
3.0 ml standard soln. 0.030
5.0 ml standard soln. 0.050
10.0 ml standard soln. 0.100
50 ml sample
mg NH3/N in 50 mix 20 = mg NHg/N per liter
13-8
-------
LABORATORY PROCEDURE FOR NITRATE
MODIFIED BRUCINE METHOD
Betty Ann Punghorst*
I REAGENTS
A Stock standard nitrate (100 mg/1 N) 0.7218
gm KN03/1.
B Working standard (1 ml = 0. 001 mg N).
Dilute 10 ml of stock to 1 liter.
C 30% NaCl solution.
D Sulfuric acid solution
Add 500 ml cone. H SO to 125 ml HO.
Cool and keep tightly stoppered.
E Brucine - sulfanilic acid reagent
Dissolve 1 gm brucine sulfate _and_ 0. 1 gm
' I.-
of
3 ml of cone. HC1. Cool. Make up
to 100 ml. Solution is stable for^several
months.
II PROCEDURE
A Preparation of Standards and Samples
1 Standards
Pipette 1.0, 2.0, 5.0, 7.0, and 10.0
ml of working standard into 1" colori-
metric tubes, held in a suitable rack
in a cold water bath. Make up to 10 ml
with distilled water.
2 Samples
Pipet 10 ml of sample (0. 5 - 8. 0 jig N)
into a 1" colorimetric tube. Run dupli-
cate analyses on each sample.
B Addition of 30% Sodium Chloride
Add 2 ml of 30% NaCl to each tube (Use
volumetric pipette). Mix well by swirling.
NOTES
1" colorimetric tubes should be matched.
Beer's Law is followed up to 10 ^g
NO,N.
O
~> \
a
*Chemist, DWS&PC Training Activities, SEC. Reviewed December 1965.
CH. N.lab.2a. 12.65
13-9
-------
Lab. Procedure for Nitrate Modified Brucine Method
Allow contents of tubes to reach tempera-
ture of cold water bath.
C Addition of Sulfuric Acid Solution
Pipet 10 ml of tL^SO^ solution into each
tube. (Use volumetric pipette) Mix well
by swirling. Allow contents to reach
thermal equilibrium in cold water bath.
D Color Development
1 Add 0. 5 ml of brucine-sulfanilic acid
solution and mix thoroughly.
2 Place tubes in boiling water bath for
20 minutes.
3 Remove tubes from boiling water bath
and immerse them in a cold water bath
and bring temperature to between 15-
25°C.
4 Adjust Spectronic 20 (410 m|i) to 100%
TRANSMITTANCE using distilled water
as a blank. Read and record % TRANS-
MITTANCE data for samples and
standards.
E Calculations
1 Plot % TRANSMITTANCE vs standard
concentration on semi-log paper.
2 Calculate concentration of nitrate
nitrogen in 10 ml aliquot of sample from
the standard curve.
3 Calculate mg/1 of nitrate nitrogen in
sample. MGNO3N in 10 ml aliquot X 100 -
mg/1 NO3N.
REFERENCES
NOTES
It is essential for good color development
that the samples and standards be mixed
well.
It is essential that the tubes be cooled
(15 - 25°C) before adding the brucine sul-
fanilic acid reagent. The color develop-
ment depends upon controlled temperature
conditions.
When samples have visible turbidity and
color, it is necessary to use the following
procedure. Before reading % transmit-
tance of the samples, adjust the Spectronic
20 to 100% transmittance using a sample
blank. The sample blank, containing all
reagents except the brucine - sulfanilic
acid solution, can be prepared while the
samples are in the boiling water bath.
The color developed is stable for 30
minutes.
Finger, James H. Nitrate Determination in
Saline and Estuarine Waters: Comparison
of Hydrazine Reduction and Brucine
Modification Methods. Laboratory In-
vestigations Report No. 3. TA&I Section.
Robert A. Taft Sanitary Engineering
Center. Cincinnati, Ohio. October, 1964.
Jenkins, David and Medsker, Lloyd L.
Brucine Method for the Determination
of Nitrate in Ocean, Estuarine and
Fresh Waters. Anal. Chem. 36:610-
612. March, 1964.
13-10
-------
DETERMINATION OF CHLORIDE AND SULFATE IN WATER SUPPLIES
D. G. Ballinger*
CHLORIDE
I OCCURRENCE
In fresh waters, high coloride values may
indicate the presence of animal pollution.
However, the chloride test as an indication
of animal pollution should be confirmed by
bacteriological and sanitary analyses. A
high chloride value can be due to other sources
such as oil field brines and other industrial
wastes; also from the passage of water through
a natural salt formation; or from agricultural
return wastes.
B Mohr
The neutral or weakly alkaline sample is
treated with chromate indicator and
titrated with silver nitrate. Silver chloride
precipitates and at the endpoint, red silver
chromate is formed. Iodide and bromide
register as equivalent chloride. Phosphate,
sulfide and cyanide interfere. Sulfite inter-
feres but can be removed with hydrogen
peroxide. Color, if present, can be re-
moved with aluminum hydroxide suspension.
II SIGNIFICANCE
Chloride compounds may break down,
especially under boiler pressures, to form
HC1 thus causing corrosion problems.
Chlorides are undesirable in ice making, as
they spoil the appearance of the ice. Approxi-
mately 500 mg/1 salt imparts an undesirable
taste to drinking water. For brewing or soft
drinks, the salt content should not exceed
275 mg/1 as the concentration may be in-
creased in the process. The U.S. Public
Health Service Drinking Water Standards for
potable waters recommend a maximum chloride
content of 250 mg/1, because of taste effects.
Ill ANALYTICAL METHODS
A Volhard
An excess of silver nitrate is added to the
acidified sample to precipitate chlorides.
The excess silver is titrated with thio-
cyanate in presence of ferric ion and
nitrobenzene. At the endpoint, red ferric
thiocyanate is formed. Iodide and bromide
register as equivalent chloride. Phosphate
and sulfite do not interfere, but sulfide
does.
C Mercuric Nitrate
The sample, adjusted to pH 3. 1*, is
titrated with mercuric nitrate solution.
Since slightly dissociated mercuric chloride
is formed, no precipitation occurs. At the
endpoint, the excess mercuric ions produce
a violet color with diphenylcarbazone in-
dicator. Bromphenol blue is added to the
indicator solution for pH adjustment. It
improves the sharpness of the endpoint by
masking the pale color produced by di-
phenylcarbazone during the titration.
Iodide and bromide register as equivalent
chloride. Sulfite in concentrations greater
than 10 mg/1 interferes but can be removed
with hydrogen peroxide. Chromate and
ferric ions interfere when in excess of
10 mg/1.
*Do not use hydrochloric acid to adjust pH
in this determination.
*In Charge, Chemistry, Technical Advisory and Investigations Section, DWSPC, SEC.
Reviewed December 1965.
CH.HAL. cl. 6c. 11.64
14-1
-------
Chloride
IV PRECISION REFERENCES
The Analytical Reference Service Mineral l Standard Methods for the Examination of
Study of 1961 reported on a reference sample Water and Wastewater. llth Edition.
containing 241 mg/1 Cl". Forty-two labora- APHA, AWWA, WPCF. 1960.
tories employed the Mohr titration procedure,
with a standard deviation of + 10 mg/1. Nine 2 Water Mineral Study of 1961. Analytical
laboratories used the Mercuric Nitrate method Reference Service, SEC.
and obtained slightly better results with a
standard deviation of +_ 7. 8 mg/1.'^'
14-2
-------
SULFATE
D. G. Ballinger*
I DEFINITION AND OCCURRENCE
Sulfate is found in all fresh waters as a
result of solvent action of water on gypsum
(CaSO4' 2H2O) and other common minerals
such as Epsom salt (MgSO4- 7H2O). Sulfates
also occur as the final oxidized stage of
sulfides, sulfites, and thiosulfates. They may
also occur as the oxidized state of organic
matter in the sulfur cycle, and, in turn serve
as sources of energy for sulfate-reducing
bacteria; at lower pH these will form hydro-
gen sulfide, which is quite undesirable.
Indusrial ,
pulp miiJ^a^nneries, ^fiifiklijJg .operations
"aHToth^r plants that use^sulfates^pr^sulfuric,
acul, contribute to the natural sulfate content
of_rawjwatejs- Sulfuric acid is the heaviest
tonnage chernicalrnanufacTuT'e'S. ""
II SIGNIFICANCE
PHS drinking water standards call for not
more than 250 mg/1 of SO4. Public water
supplies with high sulfate content are com-
monly used with no adverse effects, and this
limit does not appear to be based on tests or
physiological effects other than a laxative
action for new users. The taste threshold of
magnesium fulfate is 400 - 600 mg/1, and
for calcium sulfate is reported to be 250 -
900 mg/1. Excessive concentrations (1000 -
2000 mg/1) of magnesium sulfate may have
purgative effects.
Sulfates may be either beneficial or detri-
mental in water used for manufacturing. In
the brewing industry, the presence of sulfate
is advantageous, as it aids in producing
desirable flavor. On the other hand, Sulfates
are undesirable in the ice industry because
of the formation of white butts. In domestic
water systems, Sulfates do not appear to
cause any increased corrosion on brass fit-
tings, but concentrations above 200 mg/1 do
increase the amount of lead dissolved from
lead pipes.
Calcium sulfate scale is normally not en-
countered in once-through cooling water
systems since it is quite soluble at the
temperatures usually existing. Inrecircula-
ting systems, where concentration takes
place, the sulfate content of the circulating
water may become high enough to precipitate
calcium sulfate in the form of gypsum
(CaSO4-2H2O).
Treatment measures for preventing scale
formation are directed primarily at calcium
carbonate precipitation since it is less
soluble than the sulfate. The usual treat-
ment uses sulfuric acid which increases the
sulfate content. This, under certain
conditions may create the additional problem
of calcium sulfate scale formation.
The precipitation of calcium sulfate can be
hindered by surface active agents such as the
polyphosphates and organics. If necessary,
sulfates can be removed by evaporation, de-
mineralization, or precipitation with barium
salts.
The publication "Water Quality Criteria" of
the California State Water Pollution Control
Board, lists the recommended limits on
sulfate in mg/1, shown below in Table 1.
Ill ANALYTICAL METHODS
The gravimetric method is recognized as the
standard procedure and is the most accurate
and most time-consuming. It should be used
for sulfate in greater concentration than 60
mg/1. The turbidimetric procedure is rapid
and more accurate for concentrations less
than 50 mg/1, but can be used up to 60 mg/1.
The most rapid method is the titrimetri'c,
which is applicable to solutions containing
100 mg/1 SO4 or more, where an accuracy of
+ 10% is acceptable, as in boiler water
analysis. This procedure is not applicable to
Basic Data Network samples. Obviously,
dilution or concentration of the sample will
bring most waters into the desired range for
any of the methods.
*In Charge, Chemistry, Technical Advisory and Investigations Section, DWSPC, SEC.
Reviewed December, 1965.
CH.SUL. Ib. 11.64
14-3
-------
Sulfate
Table 1. RECOMMENDED LIMITS ON SULFATE CONTENT OF INDUSTRIAL WATER
Industrial process
Brewing
Carbonated Beverages
Concrete Corrosion
Ice Making
Milk Industry
Photographic Process
Sugar Making
Textiles
Milligrams per liter
804
--
250
25
--
60
--
20
100
CaSO4
100 - 500
300
100
—
—
MgS04
100 - 200
130 - 300
Na2SO4
100
300
A Gravimetric Procedure
The gravimetric procedure involves the
addition of a dilute solution of barium
chloride to the sample to precipitate
barium sulfate. The precipitation is made
in a solution slightly acidified with HC1
and near the boiling temperature. The
precipitate is filtered off, washed with
water until free of chloride ions, ignited
at 800°C, and weighed as barium sulfate.
= _ mg BaSO4 X 411.5
4 mi Of sample
B Turbidimetric Method
In the more rapid turbidimetric method,
sulfate ion is precipitated with barium
ion in acid solution in such manner as to
form barium sulfate crystals of uniform
size. No other ions are found in normal
waters that will precipitate with barium
in acid solution. Light transmitted by the
turbid solution is measured with a
photometer and the sulfate ion concentra-
tion is read from a standard curve. Color
and turbidity must be removed first. The
procedure described in Standard Methods
involves very careful control of stirring
and the time interval before reading. A
modification, used by the Analytical
Reference Service Laboratory at SEC
has shown much more consistent results.
IV PRECISION AND ACCURACY
The ARS Water Mineral Study of 1961
reported, on_a reference sample containing
259 mg/1 804 , that the gravimetric
procedure remains the most commonly used,
with the turbidimetric method next.
The standard deviation for the gravimetric
method was j^ 11. 9 mg/1, and for the
turbidimetric method it was 23.9 mg/1.
REFERENCES
1 Standard Methods for the Examination of
Water and Wastewater. (llth edition)
APHA, AWWA, WPCF. 1960.
2 Water Mineral Study of 1961. Analytical
Reference Service. SEC.
14-4
-------
Sulfate
3 Water Quality Criteria. State Water 4 Public Health Service Interstate Quaran-
Pollution Control Board. Sacramento, tine Drinking Water Standards
California. 1963. (revised) 1961. Federal Register,
pages 6737 - 6740. July 27, 1961.
14-5
-------
ALKALINITY AND RELATIONSHIPS BETWEEN THE
VARIOUS TYPES OF ALKALINITIES
Robert C. Kroner*
I PRELIMINARY
The property of water referred to as alkalinity
is usually caused by the presence of hydroxyl,
carbonate and bicarbonate ions. To a lesser
extent borates, phosphates and silicates con-
tribute but are generally present in negligible
amounts.
The concentration and ratio of the OH"
and HCOg" ions may be measured by titrating
a sample to certain specified pH's or end
points which are detected either by use of a
pH meter or by color indicators. Phenol-
phthalein is used for visual detection of the
first end point, (approximately pH 8) which
indicates the neutralization of NaOH and con-
version of COs to HCO3~. A number of in-
dicators (methyl orange, methyl purple, brom
cresol green etc. ) are used for detection of
the second end point (pH 3-5) which indicates
the complete conversion of HCOs" to H2O
and CO2- The final end point is determined
by the amount of COs"~ and HCOs" originally
present in the sample. If the end points are
determined electrometrically they are taken
as the mid-point of the greatest rate of pH
change per unit volume of titrant.
II RELATIONSHIPS BETWEEN HYDROXIDE,
CARBONATE, AND BICARBONATE ALKA-
LINITIES
The results obtained from phenolphthalein
and total alkalinity measurements offer a
means of classification of the principal
forms of alkalinity, if certain assumptions
are made. It must first be assumed that
interferences are absent and that bicar-
bonate and hydroxide do not exist in the
same solution. According to the system
presented in Standard Methods, llth Edition:
A Hydroxide alkalinity is present if the phe-
nolphthalein alkalinity is more than one-
half the total alkalinity.
B Carbonate alkalinity is present if the phe-
nolphthalein alkalinity is not zero but is
less than the total alkalinity.
C Bicarbonate alkalinity is present if the
phenolphthalein alkalinity is less than
one-half the total alkalinity.
Table 1. Relationships Between Phenolphthalein Alkalinity, Total Alkalinity,
Carbonate Alkalinity, Bicarbonate Alkalinity and Hydroxide Alkalinity
Lecture
Notes
Case 1
Case 2
Case 3
Case 4
Case 5
Result of
Titration
P = T
P = YiT
P = O
P > ^T
p < y2T
OH' Alkalinity
as CaCOs
T
0
O
2P-T
O
COs" Alkalinity
as CaCOs
O
2P
O
2(T-P)
2P
HCOs'Alkalinity
as CaCOs
O
O
T
O
T-2P
P = Phenolphthalein Alkalinity
T = Total Alkalinity
*In Charge, General Laboratory Services, Water Pollution Surveillance System, SEC, and re-
vised by J.\V. Mandia, Chemist, DWSPC Training Activities, SEC. Reviewed December 1965.
CH.ALK. 2a. 11.64
14-6
-------
Alkalinity and Relationships between the Various Types of Alkalinities
Table 2. Stoichiometric Volumes of Solutions of Different Normalities
Standard Solution
Normality
Equivalent Vol-
umes, ml
H2SO4
0.0200
10.0
9.4
10.0
6. 3
NaOH
0.0189
10. 6
10.0
10.5
6.6
Na2CO3
0. 0199
9.9
9.5
10.0
6. 3
NaHCOs
0. 0125
16. 0
15. 1
15. 9
10.0
III CASE EXAMPLES
The relationships involved in Table 1 may
best be explained by reference to the follow-
ing graphs. These were prepared by titrating
volumes of standard solutions of sodium hy-
droxide, sodium carbonate, and sodium bi-
carbonate with standard sulfuric acid. The
stoichiometric volumes of the various solu-
tions are summarized in Table 2 for conven-
ience in the interpretation of the charts.
A CASE 1 - Where phenolphthalein alkalinity
= total alkalinity
pH
P) END POINT
ML 0. Olfl'l N NfiOH vs 0.020 N HSO
The sharp break occurs at the point where
all of the NaOH has been exactly neutral-
ized by the acid. The pH and concentration
of the end products (Na2SO4 and H2O) de-
termine the pH at the equivalence point
between NaOH and H2SO4; in this case,
approximately 7. 0.
B CASE 2 - Where phenolphthalein alkalinity
= one-half the total alkalinity
I 1 1 1 1 1
I I I I i
5 10 IS 2(1 2r,
MI, o.o2ti N n.,so4 AIWKH
21) Ml, 0.020 N Nn2 CO, vs 0 (12(1 N H2SC>4
(P = ! T)
The titration proceeds in 2 stages where-
in all of the CO3~~ is converted, first to
HCO3-and finally to H2CO3. The first
end point occurs at approximately pH 8,
and at exactly half the volume of acid used
for the total titration. The end point
which occurs at approximately pH 4 re-
presents the total alkalinity and requires
exactly twice the volume of acid used for
the first end point.
If either HCOs" or OH" ions had been pre-
sent the titration volumes for the curves
would not have been of equal magnitude.
14-7
-------
Alkalinity and Relationships between the Various Types of Alkalinities
C CASE 3 - Where phenolphtahlein alkalinity
= 0
-(I1) KND POINT
I
J_
I
J_
J_
11) J5 20 25 i(
Ml, 0.020 N H^.SO ADDED
25 Ml. 0.0125 N NnllCO. vs, 0.020 N II_SO
,J 24
(P • O)
T The reaction proceeds in one stage with
the initial pH at approximately 8. 5 and
final pH at 4. 0. In this case the phenol-
phthalein alkalinity is zero and since no
conversion of 003"~ to HCC>3 is noted the
total alkalinity can only be due to the
HCOs" ion.
D CASE 4 - Where phenolphthalein alkalinity
is greater than one-half the total alkalinity.
ML 0.020 N 1USO, ADDED
MO MI. 0.
{11HO N N.iOII t 10 ML o.<
(P > j II
120 N N.i CO ~|
The volume of acid required for the first
end point (phenolphthalein alkalinity) is
due to the OH" neutralization and conver-
sion of the COs" to HCOs". The second
end point represents the complete conver-
sion of HCOs" to H2COs. Referring to
Case 2 where the volume of acid was
similar for each end point, it is apparent
that a base responding to phenolphthalein
but which is not COs"" must be present.
Since it was originally assumed that OH"
and HCOs" do not exist in the same solu-
tion we must conclude that the total alka-
linity is due to OH" and CO3~~.
E CASE 5 - Where phenolphthalein alkalinity
is less than one half of the total alkalinity.
Ml, 0 02(1 N H SO M)l)l I*
2 'I
FlO ML 0 020 N Nn.CO I 10 Ml, II m N N>II(O~]
If, in the reaction NaOH + NaHCO3 —»•
Na2COs + H2O, the NaOH exists in ex-
cess quantity, the final sample contains
NaOH and Na2COs, (Case 4) in which the
volume of acid required for the phenol-
phthalein end point is greater than one-
half the total. In this case, however,
the situation is reversed, wherein the
volume of acid required for the HCOs"
end point is greater than one-half the
total. Referring again to the reaction
NaOH + NaHCO3 —»• Na2COs + H2O, if
NaHCOs is in excess the end products
must consist of Na2COs and NaHCOs
and OH" must be absent. The end points
consist, therefore, of Na2COs —•
NaHCOs (phenolphthalein end point) and
NaHCO3 —>
14-8
-------
Alkalinity and Relationships between the Various Types of Alkalinlties
F CASE 6 - Where phenolphthalein alkalinity
is greater than one-half total alkalinity.
15 20 2f> JO
ML 0.020 N H..SO ADDED
QlO ML 0.0181 N NaOl'l + 10 ML 0,0125 N NnllCO ~| •
0.020 N 1LSO, ' ~*
<• 4
(p > IT}
IV
, (10 +12.6) . , ., . ,
and - - 5 - • = volume of acid required
&
for conversion of CO3~~ to HCO3~. The
j -i • . ,
second end point occurs at
(10 + 12.6)
,
ml,
the volume of HCO3~ which is converted
to H2CO3. This then becomes the same
as Case 4.
COMPARISON OF ANALYTICAL METHODS
FOR ALKALINITY (According to the
Analytical Reference Service Report
JAWWA Vol. 55, No. 5, 1963.)
Following the original assumption that OH"
and HCO3~ are not compatible, with HCO3~
being converted to CO3~~ we have a con-
dition similar to Case 4 (P>l/£r).
G CASE 7 - Where phenolphthalein alkalinity
is greater than one-half total alkalinity.
pH
~1 1 1
-(P) KNI) POINT
r-(T) KND POINT
DETEKMINATION METHODS
TABLE 1—Statistical Summary (conld.)
Method
Ye«r
No of
Vlluei
Reported
Concn.
Added
mj/l
Concn. Determined
iff /I
Mean
Low Hlih
50%
Ranie
«!/'
Sundard
Deviation
"I/I
Alkalinity
Methyl orange
Electronic trie
Methyl purple
Mixed indicator
Brom cresol green
All methods
1956
1958
1961
1956
1958
1961
1956
1958
1961
1958
1961
1958
1956
1958
1961
38
19
27
14
53
88
8
10
18
4
15
1
60
92
178
19
17
42.5
19
17
42.5
19
17
42.5
17
42.5
17
19
17
42.5
200
195
43.7
19.7
19.4
44.2
14.7
19.3
44.8
19.5
42.8
19
19.2
19.6
43.9
11.23
16
38
19.0
15
38.9
14.0
18
40
18
39
19
11 23
15
25
30.0
24
50
21.0
25
57
15.2
21
50
20
49
19
300
25
57
± 2
± 1
± 1.5
± 1
± 2
± 2.5
± 4
± 2
± 1.5
± 3
± 3
± 2
± 2
± 2
± 1.9
5.049
2483
3250
0.911
2 740
3.605
0.475
0.823
3081
1.000
3550
4460
2320
5.335
[lOML 0.0180 N NaOIl + 10 ML 0.020 N NaCOj H 10 ML
0.012r> N NaHrO(]vs 0.020 N II,,SO4
; T)
The first end point occurs at the stoichio-
metric sum of the equivalent volumes as
follows:
(9.4 - 6. 3) +
(10 + 12.6)
Alkalinity - The methods for alkalinity
measurement varied only in the choice of in-
dicator or pH for determining the endpoint of
the titration. The indicators used included
methyl orange, methyl purple, and mixed
indicator. The data show that as the use of
electrometric endpoint increased, the use of
methyl orange decreased.
where (9. 4-6.3)= volume of N acid re-
quired for excess OH" after OH" +
reaction,
14-9
-------
ALKALINITY LABORATORY
J. W. Mandia*
I REAGENTS AND EQUIPMENT
A Reagents
1 0.02NH2SO4
2 Phenolphthalein indicator
3 Methyl orange indicator
4 Mixed indicator (Brom Cresol Green-
Methyl Red)
5 Alkalinity test sample
B Equipment
1 Potentiometer
2 50 ml burette
3 Medicine dropper
4 250 ml beaker
5 Magnestir and rod
6 50 ml volumetric pipette
7 50 ml graduate
8 Stirring rod
II PROCEDURE
A Phenolphthalein-Methyl Orange
Determination
1 Pipette 50 ml of sample into 250 ml
/ beaker; add 50 ml distilled water.
^^3
•S ^2 Add phenolphthalein and titrate to
colorless end point.
3 Add methyl orange (4 drops) titrate to
yellowish-gold end point.
4 Record mis of 0.02N H?SO used for
each end point and make five
determinations.
5 Compute average titration values for
phenolphthalein alkalinity and calculate
standard deviation.
6 Compute methyl orange or total
alkalinity for each determination using
the following formula:
Total Alkalinity as
CaC03 mg/1
Ml H0SO X N X 50 X 1000
^ 4
Mis sample
7 Compute average value for total
alkalinity and calculate standard
deviation.
• — — -*
B Phenolphthalein-Mixed Indicator
Determination
1 Proceed as outlined above for
phenolphthalein end point determina-
tion (II-A).
2 Add 4 drops of mixed indicator titrate
to a grey end point pH 4. 8. This end
point is used for alkalinities ranging
from 50 - 200 mg/1.
3 Titrate to pink end point pH 4. 6; make
five determinations.
4 Compute average titration values for
phenolphthalein alkalinity and calculate
standard deviation.
5 Compute average titration values for
mixed indicator GREY end point and
calculate standard deviation.
6 Compute average value for total ,
alkalinity and calculate standard devi-
ation for mixed indicator PINK end
point. Compute total alkalinity using
the above formula.
*Chemist, DWSPC Training Activities, SEC
CH.ALK. lab. la. 12.65
14-10
-------
Alkalinity Laboratory
C Electrometric Titration
1 Place 50 ml of sample and 50 ml of
distilled water into a 250 ml beaker.
2 Place magnestir rod in beaker, lift
pH electrodes from holder and put
beaker on magnestir.
3 Insert electrode, adjust rotation of rod
to a moderate speed.
4 Push down on meter button "pH" and
titrate with rapid drops to pH 9.5 then
slowly to pH 8. 3.
5 Record mis of titrant used.
6 Titrate with rapid drops to pH 5.5,
then add drops slowly until pH 4. 5 is
reached. Record mis of titrant used.
7 Remove burette to opposite position
on clamp.
8 Push down on meter "Stand by" button.
\
9 Lift electrode and rinse into beaker.
10 Pour out sample from beaker, catch
magnestir rod and rinse.
11 Fill burette with acid and return to
position for titration.
12 Compute pH 4.5 and pH 8. 3 average
titration values and calculate standard
deviation.
13 Compute pH 4.5 total alkalinity using
above formula for each determination
and calculate standard deviation.
14 Make five determinations.
•t-
14-11
-------
Alkalinity Laboratory
o
• l-l
^
I
"o
0)
l-l
H
00
oo
Ar\
( > '
PH
(1) W
So)
h
TS °
-------
-------
DETERMINATION OF CALCIUM AND MAGNESIUM HARDNESS
B. V. Salotto*
I INTRODUCTION
A Definition of Hardness
1 USPHS - "In natural waters, hardness
is a characteristic of water which re-
presents the total concentration of
just the calcium and magnesium ions
expressed as calcium carbonate. If
present in significant amounts, other
hardness-producing metallic ions
should be included".
3 Use of water which result in change
in hardness such as:
a Irrigation
b Water softening process
B Objections to Hardness
1 Soap-destroying properties
2 Scale formation
B Other Definitions in Use
1 Some confusion exists in understanding
concept of hardness as a result of
several definitions presently used.
2 Soap hardness definition includes
hydrogen ion because it has the capacity
to precipitate soap. Present definition
excludes hydrogen ion because it is not
considered metallic. ,
"J^ ,1 *'}•>-<-""''K^-it** — t^V^y'"''"'^ TJA
3 Other agencies define hardness as "the
property attributable to presence of
alkaline-earths".
4 USPHS definition is best in relation to
objections of hardness in water.
II CAUSE OF HARDNESS IN WATERS OF
VARIOUS REGIONS OF THE U. S.
C Removal and Control
Hardness may be removed and controlled
through the use of various softening
operations such as zeolite, lime-soda,
and hot phosphate processes. It can
also be removed by simple distillation or
complex formation with surface active
agents (detergents).
Ill DETERMINATION OF TOTAL HARDNESS
A Three Methods in Use
1 Soap method
2 Compleximetric method (EDTA)
3 Calculation from individual analysis
of metallic ions other than Na and K.
A Variation
Hardness will vary throughout the country
depending on:
1 Leaching action of water traversing
over and through various types of
geological formations.
2 Discharge of industrial and domestic
wastes to water courses.
B Compleximetric Method
1 Principle of determination
Ethylenediaminetetra acetic acid (EDTA)
is a sparingly soluble amino polycar-
boxylic acid which forms slightly
ionized and very stable colorless
complexes with the alkaline-earth
metals.
*Chemist, Basic and Applied Sciences Branch, DWS&PC, SEC.
CH. HAR. 3. 12.65
15-1
<\
-------
Determination of Calcium and Magnesium Hardness
2 Interferences
Iron, manganese, nickel and zinc
interfere.
3 Procedure
Time and pH considerations.
4 Calculation of total hardness assuming
a known vol. of titrant of EDTA.
5 Precision and accuracy.
C Determination of Calcium Hardness
1 Principle of determinations
Murexide indicator forms a salmon-
colored complex with calcium whose
ionization constant is of a higher
value than that of the CaEDTA complex.
2 Interferences
Heavy metals and Sr interfere
3 Procedure
Time of titration and proper lighting
conditions are critical factors.
4 Calculation of Ca hardness.
5 Precision and accuracy.
D Determination of Magnesium Hardness
1 Calculation by difference method most
commonly used.
2 Equivalent of Ca hardness is subtrac- •
ted from total hardness equivalents, the
difference attributable to magnesium
equivalents.
3 Other methods such as pyrophosphate
method where calculation by difference
method cannot be used.
REFERENCES
1 American Public Health Assoc. , and
others. Standard Methods for the
Examination of Water, Sewage, and
Industrial Wastes. 10th Edition. 1955.
2 Ibid, llth Edition. 1960.
3 Barnard, A. J., Jr., Broad, W. C. , and
Flaschka, H. The EDTA Titration.
J. T. Baker Company. 1957.
4 U.S. Public Health Service. Drinking
Water Standards. U.S. Public Health
Service Report, Vol. 61, No. 11,
1946.
5 Rainwater, R. H. , and Thatcher, L.L.
Methods for the Collection and Analy-
sis of Water Samples. U. S. Geological
Survey Water Supply. Paper 1454.
1960.
15-2
-------
LABORATORY PROCEDURE FOR HARDNESS
J. W. Mandia*
Determination of Total Hardness, Calcium
and Magnesium Hardness
I REAGENTS
A Buffer Solution
Dissolve 16. 9 g ammonium chloride,
NH4C1 in 143 ml cone, ammonium hydrox-
ide, NH4OHPH-10.
B Na2EDTA Solution
Dissolve 4. 0 g Na2EDTA and 0. 10 g
MgCl2- 6H2O in 800 ml distilled water.
Standardize against standard calcium solu-
tion and adjust so that 1. 0 ml = 1. 00 mg
CaCOs. This usually requires the addition
of between 100 to 200 ml of distilled water.
C Standard Calcium Solution
Weigh 1. 000 g of calcium carbonate, which
has been dried overnight at 105°C, into a
500 ml Erlenmeyer flask. Add dilute
HC1 until all the CaCOs is dissolved. Add
200 ml distilled water and boil for a few
minutes to expel CO2. Cool, add a few
drops methyl red indicator, and adjust to
the intermediate orange color with dilute
NH4OH or dilute HC1 as required. Make
up to 1 liter with distilled water. 1 ml =
1. 000 mg CaCO3.
D Total Hardness Indicator
Mix 0. 4 g Eriochrome Black T plus 100 g
NaCl to prepare a dry powder mixture.
E Calcium Hardness Indicator
(Calcein) grind together in a mortar 0. 2 g
calcein (G. Frederick Smith Co.) 0. 12 g
thymolphthalein and 20 g KC1.
II PROCEDURES
A Total Hardness
1 Place 25 ml sample diluted to 50 ml
with distilled water in a 125 ml Erlen-
meyer flask.
2 Add 1 ml buffer and mix. Check pH.
If it is not pH 10, add dilute NaOH
dropwise to adjust.
3 Add indicator.
4 Titrate slowly with Na2EDTA until the
last reddish tinge disappears from the
solution, adding the last few drops
dropwise at 3 to 5 second intervals.
B Calcium Hardness
To a neutral 25 ml aliquot of sample, add
0. 2 M sodium carbonate. Add 10% sodium
hydroxide and adjust pH to 12. 5 with pH
meter. Add calcein indicator and titrate
with Na2EDTA.
Ill CALCULATIONS OF TOTAL HARDNESS
AND CALCIUM FROM RESULTS OF
COMPLEXIMETRIC DETERMINATIONS.
CALCULATION OF MAGNESIUM BY
DIFFERENCE
A Total Hardness as CaCOs
1 Assume a 25 ml water sample size and
a titrant volume of 6. 00 ml of 0. 010 M
Na2EDTA solution.
2 1. 0 ml 0. 010 M Na2EDTA solution =
1 mg
CaCO
*Chemist, DWS&PC Training Activities, SEC.
CH.HAR. lab. 1. 12. 65
15-3
-------
Laboratory Procedure for Hardness
B Calcium Hardness
1 Assume 25 ml water sample and a
titrant volume of 5. 00 ml of 0. 010 M
Na2EDTA
2 Imlof 0. 010MNa2EDTA = 0.4mgCa
1 mg CaCOg Ca
(40)
ml
CaCOgUOO)
ml
ml
= 0. 4mgCa/ml
=80mg/lCa
C Magnesium - Calculation by difference in
the same water sample.
1 Assume that the total hardness contri-
bution results only from calcium and
magnesium ions in solution.
2 Calculation of meq/1
240 mg/1 CaCO X 0. 02 meq/mg = 4. 80 meq/1
O
D Sodium Absorption Ratio
1 Sodium absorption ratio =
Na +
+
2 Soft water: 25 mg/1 total hardness as
CaCO3
200 mg/1 sodium as Na+
a 25 mg/1 X 0. 02 = 0. 50 meq/1
b 200 X 0. 0435 = 8. 70 meq/1
8. 70
.
7. 4 approximately
This is a high sodium hazard water.
4 Hard water:
100 mg/1 X 0. 02 = 2. 00 meq/1 total hardness
20 mg/1 X 0. 0435= 0. 87
This is a low sodium hazard water.
80 mg/1 Ca X 0. 0499 meq/mg = 4. 00 meq/1
3 Total hardness = 4. 80 meq/1
Calcium hardness = 4. 00 meq/1
Magnesium hardness = 0. 80 meq/1
4 0. 80meq/lMg X 12. 16 mg/meq = 9. 7 mg/1 Mg
REFERENCES
1 Standard Methods for Examination of
Water and Wastewater, llth Edition.
2 Analytical Reference Service Laboratory
Procedures.
3 U. S. Salinity Laboratory Handbook.
No. 60. 1954.
15-4
-------
LABORATORY EXERCISE FOR THE STUDY
OF VARIOUS HARDNESS INDICATORS
J. W. Mandia*
I STANDARDIZATION OF Na2EDTA
SOLUTION
A Eriochrome BJack T Indicator
1 Pipette 10 ml of standard CaCOg
solution into 250 ml beaker.
2 Add 90 ml of distilled H2O.
3 Add 2 ml of pH 10 buffer and 4 drops
of SNNaOH.
4 Add a small amount of Eriochrome
black T.
5 Titrate to a distinct blue color.
6 Make 5 determinations using Erio-
chrome black T indicator.
fi
7 Compute average Mis of titrant used, ''j.'A
8 Calculate the factor for Na2EDTA. \.&1
a 1 ml of CaCOo solution = 1 mg of
hardness as CaCOq
b 1 ml of 0. DIM Na2EDTA = 1 mg
hardness as CaCOg
c Factor for Na2EDTA = mis
CaCO3/mls N£
of
9 Compute the average factor value for
Na2EDTA
deviation.
Na2EDTA and calculate standard
10 Compute hardness values using Erio-
chrome black T indicator using
Na2EDTA standardized with this
indicator.
11 Hardness formula
„„ ,, , , Mis Na9EDTA X F X 1000
Mg/1 hardness _ ^
as CaCOg Mis sample
B Univer I Indicator
1 Proceed as described in I-A.
2 Add Univer I indicator instead of
Eriochrome black T.
3 Titrate to a distinct blue color.
4 Make 5 determinations using Univer I
indicator.
5 Calculate average factor value for
Na2EDTA using this indicator and
compute standard deviation.
C Calmagite Indicator
1 Proceed as described in I-A.
2 Add 4 drops of Calmagite.
3 Titrate to a distinct blue color.
4 Make 5 determinations using Calmagite
indicator.
5 Calculate average factor value for
NagEDTA using Calmagite indicator
and compute standard deviation.
II ANALYSIS OF HARDNESS SAMPLE
A Eriochrome Black T Indicator
1 Pipette 50 ml of hardness sample into
a 250 ml beaker.
2 Add 50 ml of distilled water.
3 Add 2 ml of pH 10 buffer and 8 drops
of SNNaOH.
4 Add a small amount of Eriochrome
black T.
*Chemist, DWSPC Training Activities, SEC.
CH. HAR.lab. 2. 12. 65
15-5
-------
Laboratory Exercise of Various Hardness Indicators
5 Titrate to a distinct blue color.
6 Make 3 determinations using this
indicator.
7 Calculate hardness using the formula:
Mis Na2EDTA X F
Mg/1 hardness _ (determined with EBT) X 1000
as CaCO3 Mis of Sample
B Univer I Indicator
1 Proceed as described in II-A.
2 Add a small amount of Univer I.
3 Titrate to a distinct blue color.
4 Make 3 determinations using this
indicator.
5 Calculate hardness using the formula:
Mis Na2EDTA X F (determined
Mg/1 hardness _ with Univer I) X 1000
as CaCO;
Mis of Sample
C Calmagite Indicator
1 Proceed as described in II-A.
2 Add 4 drops of Calmagite.
3 Titrate to a distinct blue color.
4 Make 3 determinations using this
indicator.
5 Calculate hardness using the formula:
Mis Na2EDTA X F (determined
Mg/1 hardness _ with Calmagite) X 1000
as CaCOg " Mis of Sample
III DETERMINATION OF CALCIUM
HARDNESS
A Calver I Indicator
1 Add 100 ml of hardness sample with
graduate into 250 ml beaker.
2 Add 4 drops of SNNaOH, (this adjusts
to pH 12).
3 Add a small amount of Calver I.
]
4 Titrate from pink to a distinct purple
color.
£, 5 Make 2 determinations.
6 Calculate calcium hardness
Mis Na2EDTA X F
Mg/1 Ca hardness _ (best result) X 1000
as CaCO3 ~ Mis Sample
B Calver II Indicator
1 Proceed as in III-A only add Calver II
instead of Calver I.
2 Make 2 determinations.
3 Titrate rose to blue end point.
4 Calculate calcium hardness.
C Determination of Magnesium Hardness
1 Subtract mg/1 Ca hardness as CaCOg
from total hardness as CaCOg.
2 Mg/1 Mg as CaCO3 = T.H as
CaCOg Ca.H as CaCOg
IV HARDNESS INTERFERENCE WITH IRON
A Eriochrome Black T Indicator
1 Proceed as in II-A using Iron hardness
sample.
2 Observe change in color of indicator.
It is atypical.
3 Make 3 determinations.
4 Calculate hardness.
B Univer I Indicator
1 Proceed as in II-B.
2 Observe color change.
3 Make 3 determinations.
4 Calculate hardness.
15-6
-------
Laboratory Exercise of Various Hardness Indicators
C Calmagite Indicator
1 Proceed as in II-C.
2 Observe change in color.
3 Make 3 determinations.
4 Calculate hardness.
REFERENCES
1 Standard Methods for Examination of
Water & Waste Water, 1960.
2 Water and Sewage Analysis Procedures
Cat. #9, Hack Chemical Co.
15-7
-------
Laboratory Exercise of Various Hardness Indicators
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-------
FLAME PHOTOMETRY
Robert C. Kroner*
I PRELIMINARY
Flame photometry is the art and science of
applying thermal energy (heat) to elements
in order to effect orbital shifts which produce
measureable characteristic radiations. The
color of the emission and the intensity of
brightness of emission permit both qualitative
and quantitative identification.
The application of a very hot flame (2000°C
or more) produces excitation of the element,
caused by the raising of an electron to a
higher energy level and is followed by the
loss of a small amount of energy in form of
radiant energy as the electron falls back into
its original position or to a lower energy level.
II INSTRUMENTATION
The six essential parts of a flame photo-
meter are: the pressure regulators and
flow meters for the fuel gases, the atomizer,
the burner, the optical system, the photo-
sensitive detector, and an instrument for
indicating or recording output of the detector.
These components are schematically shown
in Figure 1.
A The Atomizer and Burner
Numerous variations in atomizer and
burner design have been used. Figure 2
depicts the integral aspirator-burner used
in Beckman instruments.
YELLOW
COLLIMATING
MIRROR
PRISM BLUff-SLIT
GREEN
METER
SAMPLE
ATOMIZER BURNER
FIGURE 1. SIMPLIFIED DIAGRAM OF A FLAME PHOTOMETER
*In Charge, General Laboratory Services, Water Pollution Surveillance System, DWS&PC, SEC,
and revised by R. J. Lishka, Chemist, Analytical Reference Service, Training Program, SEC.
Reviewed December 1965.
CH. MET. 16c.5.65
16-1
-------
Flame Photometry
•Fuel
' Oxygen
Sample
Figure 2. DETAILED DIAGRAM OF
BURNER-ATOMIZER OL
The sample is introduced through the inner-
most concentric tube, a vertical palladium
capillary. A concentric channel provides
oxygen, and its tip is constricted to form
an orifice. Oxygen is passed from this
orifice causing the sample solution to be
drawn up to the tip of the inner capillary.
There the liquid is sheared off and dis-
persed into droplets. All droplets are
introduced directly into flame, with a
sample consumption of 1 to 2 ml/min.
The main requirement of the burner is
production of a steady flame when supplied
with fuel and oxygen or air at constant
pressures. In the Beckman aspirator-
burner, a concentric channel provides
oxygen to operate the atomizer and the
flame. The additional concentric channel
provides fuel for the flame.
B Optical System, Photosensitive Detector,
and Amplifier
The optical system must collect the light
from the steadiest part of the flame, render
it monochromatic with a prism, grating or
filters, and then focus it onto the photo-
sensitive surface of the detector. Use of
filter photometers is least desirable due
to their limited resolution. Flame spec-
trophotometers improve application
III
as they will separate emissions in a mix-*
ture of metals, such as the manganese
lines at 403. 3 mjj. and the potassium lines
at 404. 6 m^. Placement of a concave
mirror behind the flame so that the flame
is at the center of curvature, increases
intensity of flame emission by factor of
two.
Any photosensitive device may be used in
a flame photometer. The detector must
have a response in the portion of the spec-
trum to be used and have good sensitivity.
The photomultiplier tube is the preferred
detector for flame spectrophotometers.
The amplifier increases the signal from
the phototube and improves resolution
between close spectral lines. It also
permits identification of elements pre-
sent in samples when the concentration
is very small.
APPLICATIONS OF FLAME PHOTO-
METRY TO WATER ANALYSES
Measurement of sodium and potassium in the
past has been confined to complex, tedious,
and time-consuming gravimetric procedures.
The flame technique enables the analyst to
perform these determinations in a matter of
seconds. If these metals alone were the
only elements capable of measurement by
flame photometry the use of the instrument
could still be justified in a great many
laboratories.
Other cations which may be detected and
measured in waters and waste materials
are calcium, magnesium, lithium, copper,
and others. Table 1 includes those elements
which may be measured with commercially
available equipment including ultra-violet
and photomultiplier accessories.
Table ] does not include wavebands which
occur in the infrared spectrum. Sodium, for
example, has an emission band at 819 m(j.
which is not detectable with the common
instruments.
Many other metals, including the rare earths,
can be measured using the flame technique,
but they are not included in the table because
16-2
-------
Flame Photometry
Table 1
Aluminum
Barium
Beryllium
Boron
Cadmium
Calcium
Chromium
Copper
Iron
Wavelength
484. 2
467. 2
396. 2
553. 6
493
471
510
548
521
495
9
326. 1"
228. 8*
422. 7
622
554
425.4
360?
520
324"
327?
372
386
373
Approximate
Sensitivity
mg/1
2
3
4
0. 3
0. 4
25
100
1
2
3
2
40
0.003
0. 004
0.01
0. 1
0. 1
0. 1
0.01
0.01
0. 2
0. 2
0. 3
Lead
Lithium
Magnesium
Manganese
Mercury
Potassium
Silver
Sodium
Strontium
Zinc
Wavelength
405,
368,
364'
670. 8
371,
383"
285. 2
403^
279*
561
235. 7*
766. 5
404.6,
344. T
9
338. 3,
328. 1'
589. 3,
330. 3'
460. 7
681
407. 8
213. 9*
500
Approximate
Sensitivity
mg/1
2
2
3
0.002
0. 1
0. 1
0. 2
0.01
1
2
10
0.001
0. 2
3
0.05
0. 1
0.002
1
0.02
0.01
0. 5
500
200
* = Ultra-violet spectrum
? = Doubtful detection in visible spectrum
16-3
-------
Flame Photometry
the necessity for their measurement in water
is a rare occurrence.
IV INTERFERENCES
A Spectroscopic Interferences
Energy at other wavelengths or from other
elements than those intended to be meas-
ured may reach the detector. This prob-
lem is related to the resolution of the
instrument and slit widths used.
Many of the instrumental difficulties are
related to reproducibility of the flame.
The quality and composition of the fuel
affect the constancy and temperature of
the flame which in turn influences the
energy of emission. Likewise, slight
variations in fuel pressures and ratio
affect the reproducibility of the flame with
reference to shape, temperature, back-
ground, rate of sample consumption, etc.
In some cases the temperature of the
flame is the limiting factor in whether a
metal may be determined. (The alkaline
earth metals emit radiations at "low"
temperatures whereas other metals re-
quire very "hot" flames.) Table 2 indicates
temperatures obtainable with different fuel-
oxidant mixtures.
Table 2
Approximate Temperatures of
Fuel-Oxidant Mixtures for
Flame Photometer Use
Fuel-Oxidant
Hydrogen - air
Hydrogen - oxygen
Acetylene - oxygen
Acetylene - air*
Propane - oxygen
Illuminating gas - oxygen
Cyanogen - oxygen**
Approximate
Temp. ,°C
2100
2700 - 2800
3100
2000 - 2200
2700 - 2800
2800
4900
* Undesirable because of carbon deposits
-1* Used in research problems
Emission reading of spectral lines always *
include any contribution from the flame
background emission on which the line is
superimposed. When the photometer includes
a monochromator, it is possible to read the
background radiation in the presence of the
test element. F'irst, the line + background
intensity is measured in the normal manner
at the peak or crest of the band system.
Next, the wavelength dial is rotated slowly
until emission readings decrease to a mini-
mum at a wavelength located off to one side
or the other of the emission line or band. It
is usually preferable to read the background
at a lower wavelength than the peak. Back-
ground reading is subtracted from the line
+ background reading.
Products of combustion may affect the
characteristics of the flame or may affect
the optical system by fogging or coating of
lenses and mirrors.
B Factors Related to the Composition of
the Sample
An element may be self-absorbing, a
phenomenon in which the energy of excita-,,
tion is not proportional to the concentra- '
tion of the element. As previously dis-
cussed, excitation is followed by loss of
energy in the form of radiation as the
electron falls back to its original position
or to a lower energy level. During passage
of radiant energy through the outer fringes
of the flame, this energy is subject to
absorption through collision with atoms of
its own kind present in the ground energy
level. Absorption of radiant energy weakens
the strength of the spectrum line. Using
the emission line at 589 m^i for sodium,
Figure 3 indicates that the line ceases to
be linear at 13 ppm. As the sodium con-
centration increases, the self-absorption
effects become more pronounced. Sample
dilution to permit reading on linear portion
of the curve is often practiced.
Two or more elements present in the
sample may produce radiant energy at
the same or near the same wavelength.
For instance, calcium at 423 mp. and
chromium at 425 mp. could interfere with
each other by additive affect. The cor-
rection may be to dilute out the unwanted
16-4
-------
Flame Photometry
metal or measure one of the emissions at
a different wavelength.
STANDARD CURVE FOR SODIUM
10 20
mg/1 Sodium
Figure 3
The emission energy of one element may
be enhanced or depressed by energies
from other elements. This phenomenon
( radiation interference) occurs when one
element causes another element to modify
its actual emission intensity in either a
negative or positive manner. Correction
is obtained by dilution or by controlled
interference addition.
Other types of difficulties encountered are
too numerous to list here. In general, they
may be overcome by improved instruments
(higher resolution, narrower slit openings,
optics, flame adjustment) or possibly by
special techniques.
Some inexpensive instruments, designed
for limited use, may employ illuminating
gas with air, or propane with air as a
matter of economy or convenience.
V TECHNIQUES
The following techniques are intended to
serve as examples of current procedures in
use for routine samples and for special sam-
ples where corrective procedures are
indicated.
A Emission Intensity vs. Concentration
This is the classical procedure in flame
photometry. Solutions (standards) con-
taining known concentrations of test
element are compared with an unknown
sample. This technique is applicable only
when no interference is present.
B Radiation Buffers
For measurements of alkaline earth metals
(sodium, potassium, calcium, magnesium)
radiation buffers are prepared as solutions
saturated with regard to each metal re-
spectively. A potassium buffer, for
example, is prepared by saturating dis-
tilled water with sodium, calcium, and
magnesium chloride. A calcium buffer
in turn is saturated with sodium, potassium,
and magnesium chloride.
C Preparation of Radiation Buffers
For a sodium measurement, the buffer
solution is added equally to samples and
standards so that the interferences are
alike for all readings, thereby cancelling
each other. See Table 3.
D Instrument Improvement
Potassium emits energy bands at 766, 405,
and 345 m^. The bands are at opposite
ends of the spectrum and the 405 and 345
bands are not usable in the visible spec-
trum. The 766 line also loses sensitivity
because of its proximity to the infra—red
region. Use of a red sensitive phototube
or photomultiplier, however, permit
measurement with an ordinary instrument
at concentrations as low as 0. 1 mg/1 or
less. This approach is applicable to other
elements also.
E Standard Addition
Equal volumes of the sample are added to
a series of standard solutions containing
different known quantities of test element,
all diluted to the same volume. See Table
4. Emission intensities of the resulting
solutions are then determined at the wave-
length of maximum emission and at a suit-
able point on the flame background. After
subtracting the background emission, the
resulting net emissions are plotted linear-
ly against the concentration of the incre-
ments of the standard solutions that were
mixed with the unknown. The % transmission
16-5
-------
Flame Photometry
Table 3
Sodium Buffer
Potassium "
Calcium "
Magnesium "
NaCl
-
+
+
+
KC1
+
-
+
+
CaCl2
+
+
-
+
MgCl2
+
+
+
-
Table 4
Cone, of standards
Vol. of standard added
to sample
Vol. of sample used
Concentration of element in
each portion of mixture
0.0 mg/1
10.0 ml
10 ml
|+0 mg/1
5.0 mg/1
10. 0 ml
10 ml
|+2.5 mg/1
10 mg/1
10. 0 ml
10 ml
| + 5 mg/1
of the mixture containing unknown sample
and zero standard (distilled water) is
doubled and the concentration correspond-
ing to this point on the graph will be the
concentration of the undiluted unknown
sample. This can be explained algebraical-
ly in conjunction with Figure 4.
F Internal-Standard Method
The method consists of adding to each
sample and standard a fixed quantity of
internal standard element. The element
must be one not already present in sample.
Lithium is usually the internal standard
used. This method is most convenient
when using instruments having dual
detectors. The emission intensities of
standards and samples are read simul-
taneously or successively depending upon
instrumentation.
G Separation of Interferences
In cases where certain elements interfere,
they may be physically removed, or the
interference may be "blocked out" by
reading the emission at different wave-
lengths. To measure lithium, for ex-
ample, calcium, barium, and strontium
are precipitated as carbonates of the
metals. The lithium is retained in the
filtrate and measured at a wavelength of
671 m\i..
16-6
-------
Flame Photometry
s
o
en
w
•r-l
ra
rt
E-1 20 —
10 —
(f+0)
(ft2.5)
5)
Concent rat ion-mg/1
10
(|f5)
Let x = concentration of element in
unknown sample.
Then Y = % transmission of an equal
mixture of unknown sample
and zero standard, or
x 0
Y = -^ + -^ which simplifies to 2Y = x
. ' . 2Y = -5-+ 3. 5 (from the example in
Figure 4)
by substitution, x = — + 3.5
x = 7 mg/1
Figure 4
16-7
-------
Flame Photometry
BIBLIOGRAPHY
1 Kingsley, George R. and Schaffert,
Roscoe R. Direct Microdetermination
of Sodium, Potassium, and Calcium in
a Single Biological Specimen with the
Model Du Flame Spectrophotometer and
Photomultiplier Attachment. Analytical
Chemistry. 25: 1937-41. 1953.
2 Gilbert, Paul T., Jr. Flame Photometry -
New Precision in Elemental Analysis.
Industrial Laboratories. (Beckman Re-
print R-56) August, 1952.
3 Detection Limits for the Beckman Model
Du Flame Spectrophotometer. Data
Sheet 1. Beckman Publication. April
1952.
4 Baker, G. L. and Johnson, L. H. Inter-
ference of Anions on Calcium Emission
in Flame Photometry. Analytical
Chemistry. 26:465-568. 1954.
5 West, P.W., Folse, P., and Montgomery,
D. The Application of Flame Spectro-
photometry to Water Analysis. Analytical
Chemistry. 22: 667. Beckman Reprint
R-40. Model 10300. 1950.
6 Scott, R. K., Marcy, V.M., and Hronas, *
J. J. The Flame Photometer in the
Analysis of Water and Water-Formed
Deposits. ASTM Bulletin. Model 10300.
(Abs.) May, 1951. p 12.
7 Burriel, F., Marte, and Ramirez, J.
Flame Photometry. Munoz Elsevier
Publishing Company. New York. 1957.
8 Chow, T. J., and Thompson, T. G.
Standard Addition Method. Analytical
Chemistry. 27: 18-21. 1955.
TEXTS
1 Willard, H. H., Merritt, L. L., and
Dean, J. A. Instrumental Methods of
Analysis. D. Van Nostrand Company,
Inc. New York. 1958.
2 Dean, J. A. Flame Photometry. McGraw-
Hill Book Company. New York. 1960.
3 Clark, G. L. The Encyclopedia of
Spectroscopy. Reinhold Publishing
Corporation. New York. 1960.
-------
USE OF CONDUCTANCE MEASUREMENTS IN WATER ANALYSIS
D. G. Ballinger*
I THEORETICAL PRINCIPLES
A Ionic Transfer
An electrolyte in solution transfers electric
current by means of ion migration. Under
the influence of attractive forces, positive
ions (cations) migrate toward the cathode,
and negative ions (anions) migrate toward
the anode. Upon reaching the respective
electrode, the ions gain or lose charge,
thus transferring the charge to the external
circuit.
B Variables
The migration of ions, and thus the magni-
tude of the current flowing, is a function
of a number of variables.
1 Magnitude of EMF - The current flow-
ing through the system is directly pro-
portional to the external current applied.
2 Ion mobility - The ability of the ions
to carry the current depends on the
charge on the ion and on its size. The
common ions show a wide variability
in ion mobility.
3 Temperature - In common with most
electronic transfers, the migration
current is directly proportional to
the temperature of the solution. The
mobility of most ions increases ap-
proximately 2% for a 1° increase in
temperature.
4 Electrode area - Since a larger elec-
trode surface provides greater space
for the release or acceptance of charge,
the current is directly proportional to
the electrode area.
5 Distance - The magnitude of the cur-
rent depends on the distance ions must
travel before reaching the electrode.
6 Number of ions - When a current is
flowing through an electrolyte, each
ion carries the same amount of cur-
rent. Therefore, the greater the
number of ions present, the greater
will be the current transferred. Fur-
ther, the total current flowing is the
sum of the currents carried by the
positive and negative ions respectively.
Although the ionic transfer is in balance,
i. e., an equal charge is carried by the
cations and anions, the current trans-
fer is independent of the nature of the
opposing ion. For example, a given
number of chloride ions will carry a
particular amount of current, regard-
less of whether the cations present are
sodium, potassium, calcium or mag-
nesium.
If all of the variables discussed above
are held constant, the electrical con-
ductance of a solution is directly pro-
portional to the concentration of electro -
lyte. This principle is utilized in the
measurement of the total ion concen-
tration of a water sample.
C Specific Conductance
The conductance of a solution is actually
measured in terms of resistance to current.
Such resistance is customarily measuredin
ohms and the conductance is expressedin re-
ciprocal ohms (•"ohms) or mhos. Because
of the variables discussedinB. , conductance
must be defined under specific conditions.
Specific Conductance is the reciprocal of
the resistance measured between two
electrodes one centimeter apart and 1
square centimeter in cross section.
Tables of the specific conductance of the
common electrolytes are available in
chemical handbooks.
*In Charge, Chemistry, Technical Advisory and Investigations Section, DWS&PC, SEC.
Reviewed December 1965.
CH.MET. 15. 12.63
17-1
-------
Use of Conductance Measurements in Water Analysis
II APPARATUS
To measure the conductance of a solution, a
special cell is constructed to act as one of
the resistances of the familiar Wheatstone
Bridge.
A CeU
Two thin platinum discs are suspended
about 1 cm. apart inside a glass shell.
A coating of finely divided platinum (plati-
num black) is electroplated on the discs
to form the electrode surface. Because
of the difficulty of preparing and main-
taining an electrode surface exactly 1 sq.
cm. in area, the cell is constructed to ap-
proximately the dimensions required and
a cell constant or factor is determined. A
dipping-type cell is shown in Figure 1.
of the conductance instrument. To pro-
vide for a wide range of conductance
measurements, a series of resistances is
included in the bridge circuit. A null
point galvanometer and slide wire are
used to balance the bridge and indicate
the resistance of the sample.
C Current
Alternating current is used to prevent
polarization of electrodes.
HI PROCEDURE
A Calibration
Before making the actual measurements
of conductance, the cell constant must be
determined. Using the identical cell and
bridge, the resistance of a standard elec-
trolyte solution is carefully measured at
a constant temperature. U.01 N KC1 is
generally used, since its resistance has
been accurately determined. The meas-
ured resistance is used in the formula
K =
0.
0014118
XR
where:
K = cell constant at 25°C
R = measured resistance
0.0014118 = specific conductance of
0.01N KC1 at 25°C
Cells are commercially available having
cell constants of approximately 0.1, 1.0,
and 10.
B Measurement
CONDUCTIVITY CELL
FIGURE 1
The cell constant and the specific conduc-
tance of the sample are measured as
follows:
B Bridge
The Wheatstone Bridge, designed for
measuring resistances, is the major part
Place 4 identical tubes of standard KC1
solution and 2 tubes of sample in a
water bath at 20° - 30°C. Allow 30
minutes for temperature equilibrium.
17-2
-------
Use of Conductance Measurements in Water Analysis
Rinse the conductivity cell in 3 of the
tubes of KC1 solution and measure the
conductance of the 4th. Record this
value as
3 Rinse the cell thoroughly in the first
tube of the sample and measure the
conductance of the second tube. Re-
cord this value as Rs.
4 Calculate the specific conductance (in
H mhos) of the sample by the formula
Sp. Cond.
1.411.8 X RKC1
Re
IV APPLICATIONS
A Total Dissolved Residue
The conductivity of natural waters is di-
rectly related to the concentration of dis-
solved solids. The measurement of con-
ductivity is very rapid, does not destroy
the sample, and has good precision.
The specific conductance of a particular
sample depends on the nature of the ions
present, so that no universal relationship
can be determined for the ratio of con-
ductance to total dissolved solids. On a
particular water, however, the corre-
lation can be determined experimentally
and used to provide a rapid, convenient
index of the total ionic concentration.
The ratio of TDS to specific conductance
for several different water samples is
shown in Table I.
Note that the specific conductance varies
directly with TDS. It is interesting, how-
ever, to note the difference in ratio of the
Arkansas River samples at two stations.
The lower ratio at Ponca City is probably
due to the increase in chlorides, as shown
in Table H.
It is apparent that the nature of the ions
present must be considered in any inter-
pretation of conductance-dissolved solids
relationships.
B Checking Mineral Analysis
Because of the relationship of specific
conductance to ionic concentration, con-
ductivity may be used to verify the results
TABLE I
Sample
Arkansas River at
Coolidge, Kansas
Arkansas River at
Ponca City, Okla.
Colorado River
Cincinnati tap
Sp. Cond. (nmhos)
3330
1920
1000
590
TDS(mg/l)
2600
1040
680
430
RATIO TDS/cond.
0.73
0.54
0.68
0.73
TABLE II
At Coolidge
At Ponca City
Chloride
80
250
Sp. Cond.
3300
1920
TDS
2600
1040
RATIO TDS/cond.
0.78
0.54
17-3
-------
Use of Conductance Measurements in Water Analysis
of mineral analyses of water samples.
Rossum^' found that a satisfactory cor-
relation could be obtained in 92% of the
samples investigated. Equivalent con-
ductance of the common ions is shown in
Table III.
TABLE HI
ION
Cl
S04
C03
HC03
N03
Ca
Mg
Na
H mhos per mg/1 (25°C)
2.14
1.54
2.82
0.71
1.15
2.60
3.82
2.16
In practice, the concentration of each ion
(mg/1) is multiplied by the appropriate
factor and the sum of the individual con-
ductances is calculated. This value
should agree with the specific conduc-
tance of the sample, within + 2%. For
best results, the sample should be di-
luted to Ca 100 (imhos with high quality
distilled water. The specific conduc-
tance of the sample is calculated from
the formula:
ADX106 ._ ,. „
'd= —R5~-
-------
Use of Conductance Measurements in Water Analysis
D Selecting Aliquot Size
In many types of analyses, aliquots of the
sample must be chosen to keep the con-
centration of reactants within definite
limits. Alter suitable correlations are
established, a quick measurement of
conductance will help in selecting the
proper aliquot.
REFERENCES
1 Harley, J.H. and Wiberley, S.E., "In-
strumental Analysis", John Wiley &
Sons, New York, 1954.
2 Scofield, C. S., "Measuring the Salinity
of Irrigation Waters and of Soil Solu-
tions with the Wheatstone Bridge",
U.S. Dept. Agr. Circ. 232, 1932.
3 Wilcox, L.V., "The Quality of Water
for Irrigation Use", U.S. Dept. Agr.
Tech. Bull. 962, 1948.
4 Wilcox, L.V., "Electrical Conductivity",
JAWWA42;775, 1950.
5 Rossum, J.R., "Conductance Method for
Checking Accuracy of Water Analysis",
Anal. Chem. 2i:631, 1949.
6 Standard Methods for the Examination of
Water and Wastewater, llth Edition,
APHA, AWWA, WPCF, 1955.
17-5
-------
PRINCIPLES OF ABSORPTION SPECTROSCOPY
Betty Ann Punghorst*
I INTRODUCTION
In any system employing principles of
absorption spectroscopy there are three
basic components:
A A SOURCE of Radiant Energy
B A MEDIUM (Sample) which Absorbs
Radiant Energy
C A DETECTOR to Measure the Radiant
Energy Transmitted by the Sample
RADIANT ENERGY
DETECTOR
Figure 1. Basic Components of Absorption
Spectroscopy System
II RADIANT ENERGY
A Wave Nature
1 The various forms of radiant energy
have been arranged in a single schematic
diagram referred to as the electro-
magnetic spectrum. (See Figure 2.)
All of the energies which make up this
spectrum may be represented graphically
as waves. All waves move through
space (and for most purposes, air) at
a constant velocity, 3 X 101" cm/sec.
2 Three variable characteristics of
individual waves serve to differentiate
each from all other waves in the
spectrum.
a The Wave Length: \, the linear
distance between the crests of two
adjacent waves. (Units: distance/
wave)
b The Frequency: v, the number of
waves which pass a given point in
a unit of time. (Units: waves/time
unit)
c The Wave Number: v The number
of waves which occur in a given
linear distance. (Units: waves/
distance unit)
3 It is evident that more waves of short
wave length will "fit" into a given
linear distance than would waves of a
greater wave length. Thus, waves
having short wave lengths will have
higher wave numbers. Mathematically
wave length is the reciprocal of wave
number, if the same units of linear
measurement are used in each
expression. Since the velocity of all
waves is equal and constant, it is also
apparent that a greater number of
waves of short wave length can pass
a given point in a unit of time than
waves having a longer wave length.
B Particle Nature
Planck conducted certain experiments '
which indicated that light has a particle
as well as a wave nature. Energy rays
can be said to consist of particles with
a definite amount of energy. These
particles or packets are referred to as
photons or quanta. The energy (E) of each
minute packet is given by Planck's equation:
*Chemist, DW3&PC Training Activities, SEC. Reviewed December 1965.
CH. MET.al. 5a. 12. 63 18-1
-------
Principles of Absorption Spectroscopy
THE ELECTROMAGNETIC SPECTRUM
GENERATOR
60 CYCLE
«t ULTRa-
Mr,. T°rS
b^, -'
V OLET
*>.
VE L ENG1 H 7 6 i 10 = c>
N
T
\ 1
0
• 1 I 1 1
5
f Trough
, , , 1 ,
10
, |
13
(1)
(2)
(3)
(4)
WAVE LENGTH
X
distance
wave
X (cm/ wave)
X
X
X
X
FREQUENCY
v =
waves
time
v (waves/ sec) =
CONSTANT
c
distance _ ,
time
C = (3X lolO
VELOCITY
cm/ sec)
Figure 3. Relationship of Wavelength and Frequency
E
Where
= hv
(5)
E = radiant energy in ergs
h = Planck's proportionality
constant (6.6 X 10~27
erg sec)
v = frequency in waves per
second
Thus it can be seen that the energy of a
given photon is directly proportional to
the frequency of the given radiant energy.
Ill ABSORPTION OF ENERGY BY ATOMS
AND MOLECULES
A Absorption of energies of given frequencies
by atoms and molecules can be used as a
18-2
-------
Principles of Absorption Spectroscopy
WAVELENGTH A°
2, 000
4,000
8,000
200.,
Galon
X-rays
es per mole
needed for change
Gf:c
Ultraviolet
1*2,000
©
n'o
T lonization
Inner
Electron Shift
Visible Ne
71,000 35,
O-o C
ar IR Far IR
000 1,400
**" -*-^
)To O:o
Outer Vibration Rotation
Electron
Shift
Figure 4. Electromagnetic Spectrum Showing Energy Ranges and
Corresponding Electronic, Vibrational and Rotational Motions
basis for their qualitative identification.
Absorption spectroscopy is based on the
principle that certain displacements of
electrons or atoms within a molecule are
permissible according to the quantum
theory. When radiant energy of the same
energy required to bring about this permis-
sible change is supplied to the molecule,
the change occurs and energy is absorbed.
1 Displacement of electrons is a permis-
sible change which can occur when
energy of ultraviolet and visible frequen-
cies strikes certain atoms and molecules.
a Inner electron shift
Electrons located in the inner orbit
of an atom may, when the proper
frequency of radiant energy is avail-
able, shift to an orbit farther re-
moved from the nucleus. This shift
represents a change from a lower
energy level to a higher one. If
this new position is unstable, the
electron may revert to some posi-
tion nearer the nucleus; the energy
which is thus gained may then be
emitted from the atom as part of its
emission spectrum. The number
of energy changes possible within
an atom is a function of the number
of electrons and the number of
changes each may enter. Each
possible change gives rise to a new
spectral frequency. Since the fre-
quency of the radiation needed to
accomplish such changes is of a high
order of magnitude, the energy used
is considerable in quantity. Molecu-
lar aggregations often disintegrate
in such circumstances; thus, these
higher frequencies are used mainly
for work with elements or very stable
compounds.
b lonization
Under a specific frequency of radia-
tion, an electron may be physically
separated from its parent atom.
This process has been termed
ionization. A change of energy
level of this magnitude requires less
energy than the inner electron shift.
Such changes are characteristic of
those of the rare earths, inorganic
ions, transition elements and many
organic compounds under frequencies
within the ultraviolet range.
c Outer electron shift
The various orbital electrons in an
atom may vary in the amount of
energy required to shift them out-
wardly from the nucleus. For
example, it requires less energy to
18-3
-------
Principles of Absorption Spectroscopy
shift an electron from a position
more distant, than it does to shift
an electron outwardly from the inner
orbit. Outer electron shifts occur
readily in colored organic molecules
for which electronic transitions are
made easier by the presence of
chnomophore groups which partici-
pate in resonance. Thus the excita-
tion of the delocalized outer electrons
(pi electrons) is relatively easy and
requires energy in the visible range.
Vibration of atoms within molecules is
a permissible change which can occur
when energy of near infrared frequency
strikes certain organic molecules.
The atoms within a molecule are held
together by attractive bonding forces.
Atoms within a molecule are constantly
moving toward and away from other
atoms, but for purposes of theory, can
be said to have a certain "average"
position. The change in position of an
atom in relation to another atom is
called vibration. The mechanics of
vibration require energy; the manner
and rate of vibration of the atoms
depend upon frequencies of electro-
magnetic radiation which strikes them.
Therefore, a specific part of a molecule
may absorb significant quantities of
certain spectral frequencies. Such
absorption will be reflected in the ab-
sorption spectrum of the compound.
The energy requirements for this type
of energy change are of a lower order
of magnitude than those above; there-
fore, we would expect that the fre-
quency required would be lower and
the wave length longer. Such changes
occur in organic compounds under
infrared radiation.
Rotation of molecules is a permissible
change which can occur when energy of
far infrared frequency strikes certain
organic molecules.
A molecule rotates around its sym-
metrical center. The manner and
rate of rotation, again, depends upon
the energy supplied to it. Specific
spectral frequencies of electromagnetic
radiation can be employed to increase
the rate of rotation. The used radia-
tion is, in effect, absorbed and is
reflected in the absorption spectrum.
Organic molecules utilize infrared
radiation while varying their rate and
manner of rotation.
B The Lambert-Beer Law provides the
basis for quantitative analysis by
absorption spectroscopy.
1 Optical density
a The decrease in intensity of energy
per unit thickness of the sample i»
proportional to the incident intensity
of the energy and the concentration
of the sample.
-dl .
db Klcl
(6)
I
dl
= intensity of incident energy
= increment of incident energy
absorbed by sample
db = increment thickness of
absorbing solution
c = concentration of absorbing
solution
K! = proportionality constant
(varies with wavelength of
light, the solvent used and
the temperature).
Integrate equation (6)
db
(7)
log — = ABSORBANCE = OPTICAL
DENSITY = ac
18-4
-------
Principles of Absorption Spectroscopy
a =
K
1
2.303
= ABSORBANCY INDEX
(8)
When c is expressed in moles/1
and b in cm,
a = MOLAR ABSORBANCY INDEX
m
BOUGUER - LAMBERT LAW:
log — = ABSORBANCE = OPTICAL
DENSITY = a2bc
2.303
Optical density is directly proportional to the
length of the path the energy must travel
within the sample.
b The decrease in intensity of energy
per unit concentration of the sample
is proportional to the incident inten-
sity of the energy and the thickness
of the sample.
BEER'S LAW:
Optical density is directly proportional to
the concentration of the solute.
2 Transmittance is the fraction of
radiant light energy which passes
through the absorbing solution.
__
dc
K2bl
(9)
T = -L = 10-abc
o
(12)
b = thickness of sample
dc = increment concentration of
absorbing solution
I = incident light energy
I = final light energy
proportionality constant (varies
with wavelength of light, the
solvent used and the tempera-
ture)
100 X -i- = PERCENT TRANSMITTANCE
o
IV SUMMARY
RANGE
Ultraviolet
Visible
Infrared
SOURCE OF
RADIANT ENERGY
Hydrogen Arc
Incandescent
Tungsten Bulb
Nernst Glower
Globar Lamp
ABSORPTION BY SAMPLE
CHEMICAL NATURE
OF SAMPLE
Inorganic Ions and
Organic Molecules
Colored Inorganic and
Organic Molecules
Organic Molecules
TYPE OF SAMPLE
CELL USED
Quartz Fluorite
Glass
Sodium Chloride or
Potassium Bromide
DETECTION OF
RADIANT ENERGY
TRANSMITTED
Photoelectric Cells
Photographic
Plates
Eye Photographic
Plates
Photoelectric Cells
Thermocouple
18-5
-------
Principles of Absorption Spectroscopy
REFERENCES 2 Mellon, M.G. Analytical Absorption
Spectroscopy. John Wiley & Sons, Inc.
1 Delahay, Paul. Instrumental Analysis. 1950.
The MacMillan Company. New York.
1957.
18-6
-------
METAL COMPLEXES AND CHELATES IN THE COLORIMETRIC
DETERMINATION OF METALS
James W. Mandia*
I METAL COORDINATION COMPOUNDS
A Metal Complexes
1 When a metal ion combines with an
electron donor, the resulting substance v.~
is said to be a complex. AJL.^ ^ r
Metal + 4A
A
A — M — A
V
A
I
2 In compounds such as I Co (NHJ
the ammonia cobalt bonds do not dis-
sociate in solution and that the ammonia
molecules are held firmly in definite
positions in space about the central
cobalt atom.
NH_
H3N
\\/
/Co-NH3
+ 3
3 Cl
It is apparent, therefore, that the bonds
within the complex are covalent in
character.
The formation of Coordination com-
plexes falls under the electronic theory
of acids and bases proposed by Lewis
(1923).
According to this concept neutralization
of an acid with a base involves the
formation of a coordinate bond.
Acid Base
Cu+2 + 4 NH
Ag+1 + 2 CN
O
"1
[cu
-------
Colorimetric Determination of Metals
Example: The aluminum lake of Alizarin
O
1—OH
D Chelate Compounds and Color
1 The color of chelate compounds is
generally accepted as being so signifi-
cant that colors are, as a rule, very
carefully described in reports of these
compounds.
2 Absorption spectra of metal chelates
are quite different from those of the
organic compound and of the metal
alone.
3 For a given chelating agent, the wave
lengths of the absorption maxima are
practically independent of the metal
employed. Color is due to the presence
of the chelate ring and not character-
istic of the particular metal in the ring.
4 The intensity of absorption however,
is quite sensitive to the nature of the
metal.
E Measurement of Intensity of Absorption -
the Molecular Extinction Coefficient
£ = Molecular Extinction Coefficient
A = Absorbance
b = length of cell, usually 1 cm
C = Concentration in moles/liter usually
1 mole/1.
T =
^ bC~
II METAL CHELATE COMPOUNDS IN THE
DETERMINATION OF COPPER
A The Ferroin Group
1 The reaction of 1,10-phenanthroline
with ferrous ion yields an intensely
red, soluble compound called "ferroin. "
2 The color change from blue to red of
the oxidation - reduction couple
Fe(l, 10-phenanthroline) 3 —
Fe(l, 10-phenanthroline) „ furnishes
a high potential oxidation-reduction
indicator of great utility.
* *
O ••-" v_<
/ \
N N
// •*•
Ferroin group
B The Cuproine Group
1 It was observed that the ferroin reaction
failed with those compounds bearing
substituent groups on the carbon atom
adjacent to the ring nitrogen atoms.
2 Compounds which failed to give the
ferroin reaction produced colors with
cuprous copper. These substances
are specific for copper and are called
the cuproine group.
\
X
\
X
Cuproine group
There are three cuproine reagents -
cuproine, neocuproine and bathocup-
roine, and they react only with cuprous
copper, Cu+l.
The copper compounds of these three
cuproine reagents are intensely colored
and are soluble in certain organic
solvents.
19-2
-------
Colorimetric Determination of Metals
5 Hydroxylamine hydrochloride is usually Bathocuproine. Published by the G.
chosen as the reducing agent to reduce Frederick Smith Chemical Co. 1958.
Cu+2 to Cu+1.
2 Standard Methods for the Examination of
Ammonium acetate as the agent for Water and Wastewater. llth Edition.
suitably buffering the solution. 1962.
3 Martell, A.E. and Calvin, M. Chemistry
REFERENCES of the Metal Chelate Compounds.
Prentice-Hall Chemistry Series.
1 Diehl, H. and Smith, F.G. The Copper 1953.
Reagents, Cuproine, Neocuproine and
19-3
-------
IRON AND MANGANESE IN WATER
Betty Ann Punghorst*
I INTRODUCTION
A Sources*1)
1 Nonacid ground waters
a Iron occurs as soluble Fe(HCO3) 2-
The colorless, clear iron-bearing
ground water develops a turbid,
reddish brown color upon standing
in air due to the oxidation of
Fe++ >• Fe+++.
b Manganese occurs as soluble
Mn(HCO3)2.
2 Acid surface waters
a Iron occurs as soluble FeSO4 usually
together with MnSO4, H2SO4 and
A12 (SO4)3. Acid waters generally
do not cloud upon standing in air
unless first neutralized.
b Manganese occurs as soluble
3 Colored waters (found in the south)
a Iron occurs in an organic chelated
form in waters where color is due to
the extraction of organic materials
from decaying vegetation. Iron in
this form does not precipitate when
the water is aerated.
b Manganese occurs in a chelated form.
4 Red waters
Iron occurs as insoluble, suspended
Fe(OH)3 where the metal of water
mains, piping and tanks has corroded.
B Effects*5^
1 Water treatment sand filters are clogged
and deposits of Fe(OH)3 and MnO2 are
formed on distribution pipes. These
deposits can break loose, be carried
down to smaller pipes or valves, and
cause clogging and new corrosion.
Industrial use of water is affected,
(e. g., pulp and paper and textile
industries).
Domestic use of water is affected by
staining plumbing fixtures, forming
deposits on laundry, and imparting
unpleasant colors and tastes to
beverages.
II DETERMINATION OF IRON
A Collection of Sample*7)
^/l Iron contamination can come from the
flaking of rust in pipes or from a metal
cap on the sample bottle.
2 Colloidal iron may adhere to the sides
of a plastic sample bottle.
B Methods
1 1, 10-phenanthroline*2'6'8)
a Reaction
1) Ferric iron is reduced to ferrous
iron with hydroxylamine.
4Fe
3+
+ 2NH2OH
4Fe +
N20 +
H_O
£t
4H
2) Three molecules of 1, 10-phenan-
throline chelate one molecule of
Fe to form an extremely stable
deep red complex, which follows
Beer's Law and exhibits maximum
absorbance at 508 m(J..
*Chemist, DWS&PC Training Activities, SEC.
CH. ME.fe. 5a. 11.64
Reviewed December 1965.
20-1
-------
Iron and Manganese in Water
Fe
(2)
b Interferences^ '
There are several classes of ions
which interfere with the 1, 10-
phenanthroline test for iron.
1) Ions which form iron complexes
(e. g., phosphate, polyphosphate,
CN~). This interference can be
eliminated in the pretreatment
of the sample by boiling with HC1.
2) Ions which form only slightly
soluble complexes with 1, 10-
phenanthroline (e. g., Bf1"^,
Ag+, Cd++, Kg"1"*", Zn++). This
interference can be reduced by
adding a large excess of 1,10-
phenanthroline.
3) Ions whose own color obscures
the phenanthroline complex color
(e.g., Ni++, Co++, Cr*4"*). This
interference can be reduced by
using a blank containing an equiva-
lent amount of the interfering ion
present in the sample.
4) Oxidizing agents which prevent
the complete conversion of
Table 1 EKPKCT OF ANIONS
(All Sampler C
ON 1, 10 -PHENANTHROLINE
ontained i mg/1 iron)
Ion
Acetate
TetraboratefabBjC^
Bi omidf.
C arbonale
Chlorate
CMoride
Citrate
Cyanide
DiUiromale
Fluoride
Lodide
Nitralt-
NitriU
Oxalate
Porehloiate
Phosphate (ab P^y
Pyrophosphate
Silicate
Sullate
Sullite
fditidte
I Inotyanali
Ihiofaullalc
NaC2HjO2
tJa2B4O?
"NaBr
Na^C03
KCIOj
NaCl
C[,H807
KCN
K2Cr2°7
N.U.
KI
KNOj
KNO2
fNiy_,C204
K('I04
(NI^2HP04
Na4P207
NajbiOj
(Niy2b04
Na^SO.,
(Niy^i^,.
KC.NS
N*r"Ps__
Maximum
•ng/1
500 0
500 0
SOO 0
500 0
500 0
1000 0
500 0
1 10 0
100 0
20 0
500 0
riOO 0
500 0
500 o
500 0
100 0
20 0
50 0
20 0
100 0
50 0
500 0
500 0
500 0
50(1 0
500 0
Maximum
% Ft-
None
None
Num.
None
None
None
None
2 0
Change hue
None
1 fa
None
None
None
None
1 2
1 4
1 0
1 0
None
None
None
None
NOIIL
Nona
None*
pll Range
20-9 0
-J 0-9 0
2090
J 0-y 0
25-90
2 0-9.0
I 0-9 0
2 0-9 0
25-90
4.0-9 0
2 0-y I)
2 0-9 0
2 5-9. 0
60-90
2 0-9 0
20-9 0
60-90
5 5-9 0
2 0-4 5
20-5 I)
2 0-9 0
20-90
1 0-9 1)
20-90
J 0-9 0
'i (inbuilttdruj detLimin<_d within
dm,ed wilti hydroxylainmu hydio
oloi d< vt lop. d ii
20-2
-------
Iron and Manganese in Water
' Table 2. EFFECT OF CATIONS ON 1, 10 -PHENANTHROLINE TEST
(All Samples Contained 2 mg/1 Iron)
Ion Added As
Aluminum A1C13
Ammonium NH4C1
Antimony SbCl3
Arsenic As2°5
Arsenic AS2O3
Barium BaCln
Beryllium Be(NO3>2
Bismuth Bi(NO3)9
Cadmium Cd(NO3)2
Calcium Ca(NO3)2
Chromium Cr (SO ),
Ct 4 O
Cobalt Co(NO )
Copper Cu(NO)
•J £
Lead Pb(C2H3°2>2
Lithium LiCl
Magnesium Mg(NO3)2
Manganese MnSO
Mercury HgC19
Mercury Hg_(NO_)_
Molybdenum (NH.J.Mo^O-.
4 o t Z4
Nickel Ni(NO,,)0
O 6
Potassium KC1
Silver AgNO
Sodium NaCl
Strontium Sr(NO )
Thorium Th(NO3>4
Tin H0SnCl .
2 6
Tin H2SnCl
Tungsten Na^WO
Uranium UOJC TTOJ0
2 2 3 22
Zinc Zn(NO3)2
Zirconium Zr(NO )
15.0 ml of o-phenanthroline in
Maximum Maximum
Concentration Interference
mg/1 % Fe
500 . 0 None
250.0 1.4
500.0 None
30.0 None
500 . 0 None
500.0 None
500.0 None
500.0 1.3
None
50.0 1.0a
500.0 None
20 . 0 None
10.0 1.5
10 . 0 None
500.0 None
500 . 0 None
500.0 None
500.0 None
1 . 0 None
10.0 None
100.0 None
2.0 None
1000.0 None
None
1000.0 None
500.0 None
250.0 1.5
20.0 None
50.0 None
10.0 None
20.0 None
10.0 Negligible
100.0 None
10.0 None
50.0 1.8
100.0 2.2
Applicable
pH Range
2.0-3.0
2.0-5.0
2.0-9.0
3.0-9.0
3.0-9.0
3.0-9.0
3.0-9.0
3.0-5.5
3.0-9.0
2.0-9.0
2.0-9.0
3.0-5. 0
2.5-4.0
2.0-9.0
2.0-9.0
2.0-9.0
2.0-9.0
2.0-9.0
3. 2-9. 0
5.5-9.0
2.5-9.0
2.0-9.0
2.0-9.0
2.0-9.0
2.0-9.0
3.0-6.0
2.5
2.0-6.0
2.0-3.0
2.5-9.0
2.0-6.0
2.0-9.0
2.0-9.0
3.0-9.0
excess of the original amount added. Iron
reduced with hydroxylamine hydrochloride in all tests.
These tables have been reproduced, with permission of Ind. and Eng.
Chemistry, 10:60 (1938), by the ~
Department of Health, Education,
and
Welfare, Public Health Service.
20-3
-------
Iron and Manganese in Water
Fe+++ —*• Fe++. This interference
can be reduced by adding a large
excess of hydroxylamine.
c Extraction method'''
Iron rnay be isolated from inter-
fering ions by extracting it from
aqueous, acid solution with di-
isopropyl ether. A subsequent
aqueous extract of the iron from the
ether is then used in the colorimetric
determination with 1, 10-phenanthroline.
2 2,2', 2" - Terpyridyl (tripyridine)*4)
Two molecules of terpyridyl chelate
one atom of Fe to form a reddish-
purple complex which obeys Beer's
Law and exhibits maximum absorbance
at 555 ran. Ethylenediamine is used to
complex all the heavy metals which
might otherwise interfere.
Ill DETERMINATION OF MANGANESE
A Collection of Sample
Acidification of sample avoids the pre-
cipitation of MnO2 and its consequent
adherence to the sides of the sample
container.
B Methods
1 Periodate^3.6)
a Reaction
1) Elimination of interferences
2) Oxidation of Mn++
2Mn + 5IO
2MnO + 5IOq + 6H
TC J
Table J COMPARISON OF EFFECT OF DIVERSE IONS ON
1, Ifl-PHENANTHKOLINK, NON-SFQUESTERED, AND
SEQUESTKRKD TEHPYHIDYL METHODS
Ion
Coupe i
Cobdll
Nukcl
Zm.
Ol lliuphnspliat,.
_>•_!" "»"«"l*
-------
Iron and Manganese in Water
e 4 LFFFC f OF VMOVS ON THK PERIODA1 h 1 F,ST
Table 5. EFFECT OF CATIONS ON THE PERIODATK TEST
(0.4933 mg of manganese in 250 ml of solution)
Ion
Aluminum
Ammonium
Antimonous
Barium
Beryllium
Bismuth
Cadmium
Calcium
Chromic
Cobaltous
Cupric
Ferric
Ferrous
Lead
Lithium
Magnesium
Mercuric
Mercurous
Mickelous
Potassium
Silver
Sodium
Stannic
Stannous
Strontium
Thorium
Uranyl
Zinc
Zirconium
Concentration
mg/250 ml
100
100
ICO
100
50
50 (20 ml H2SO4)
100
100
50
10
50
10
50
10
100 (10 ml H.PO.)
5
100 (10 ml Iljl-i',)
100
100
100 (20 ml H2SO,)
100
50
50
10
125
100 (20 ml H2SO4)
155
50
20 (20 ml H2S04)
100
100
50 (U)
100
100
Apparent Change
In Manganese
Concentration
%
Negligible
Negligible
Negligible
Negligible
Negligible
Turbidity
Negligible
Negligible
Change in hue
Change in hue
+ IB. 1
+ 30
Change in hue
Change in hue
N.. eligible
Change in hue
Negligible
Negligible
Negligible
Negligible
- 50.7
1. 1
Change in hue
Negligible
Negligible
Negligible
Negligible
Negligible
Tut bidity
Negligible
Negligible
- 2.0
Negligible
Negligible
Approximate
Limiting
Concentration
mg/250 ml
0.0
0.0
5
5
0 0
50
10
0.0
50
These tables have been reproduced, with permission of Ind. and Eng. Chemistry, 11: 274 (1939),
by the Department of Health, Education and Welfare, Public Health Service.
20-5
-------
Iron and Manganese in Water
Correction for turbidity or inter-
fering color can be made by
bleaching the permanganate color
with a reducing agent and deter-
mining the absorption of the
interferences.
4) Ions which are capable of reducing
either IOi or MnC-4 (e.g., NC>2,
C20|. Sb*+).
This interference can be elimi-
nated in the pretreatment by
boiling the sample with ^SO^
and H3PO4.
2 Persulfate^
a Reaction
1) Elimination of interferences
2) Oxidation of Mn"1"4"
2Mn++ + 5S0O^ + 8H0O »-
2MnO + 10SO" + 16H
b Interferences
The interferences in the persulfate
test and their removal correspond
to those in the periodate test. How-
ever, when using the persulfate test,
HgSC>4 can be used to complex Cl~
ion.
C Precision
The Analytical Reference Service Water
Metals Study of 1962 reported on a syn-
thetic reference sample containing
0.25 mg/1 manganese. 93 results (per-
sulfate method) had a standard deviation
of + 0.100 mg/1. 57 results (periodate
method) had a standard deviation of
+ 0. 181 mg/1.
IV SUMMARY
The U. S. Public Health Service has set the
recommended limits for iron and manganese
content in finished potable waters at 0. 3 mg/1
and 0.05 mg/1, respectively. Reliable
quantitative tests for iron and manganese in
trace amounts are important in the following
areas;
A Selecting New Sources for Water Supplies
B Determining Water Treatment Processes
C Preventing Corrosion in Pipelines
REFERENCES
1 Connelley, E.J. Removal of Iron and
Manganese. Journal of the American
Water Works Association. 50:697-702.
1958.
2 Fortune, W. B. and Mellon, M. G.
Determination of Iron With
o-Phenanthroline, A Spectrophometric
Study. Ind. and Eng. Chemistry.
Analytical Edition. 10:60. 1938.
3 Mehlig, J.P. Colorimetric Determina-
tion of Manganese With Periodate,
A Spectrophotometric Study. Ind. and
Eng. Chemistry. Analytical Edition.
11:274. 1939.
4 Morris, R.L. Determination of Iron in
Water in the Presence of Heavy Metals.
Analytical Chemistry. 24:1376-1378.
1952.
5 Riddick, Thomas M., Lindsay, Norman L.,
and Tomassi, Antonio. Iron and
Manganese in Water Supplies. Journal
of the American Water Works
Association. 50:688-696. 1958.
6 Sawyer, Clair N. Chemistry for Sanitary
Engineers. McGraw-Hill Book Co., Inc.
New York. 1960.
20-6
-------
Iron and Manganese in Water
7* Standard Methods for Examination of Water 8 Welcher, Frank J. Organic Analytical
and Wastewater. llth Ed. APHA. Reagents. D. Van Nostrand Company,
AWWA. WPCF. 1960. Inc. New York. Ill: 1947.
20-7
-------
THE DETERMINATION OF TOTAL IRON BY THE PHENANTHROLINE METHOD
D. G. Ballinger and B. A. Punghorst*
I REAGENTS
A Hydrochloric Acid, Concentrated
B Hydroxylamine Reagent, 10% Solution
Dissolve 10 g NH2OH. HC1 m 100 ml
distilled water.
C Ammonium Acetate Buffer Solution
Dissolve 250 g NH4C2H3O2 m, 150 ml
distilled water. Add 700 ml glacial acetic
acid and dilute to a liter.
D Iron Stock Solution
Add 20 ml concentrated H2SO4 to 50 ml
distilled water and dissolve 0.7022 g
Fe(NH4)2 (SO4)2- 6H2O. Add dropwise
0. IN KMnO4 until a faint pink color
persists. Dilute to a liter. This solution
contains 0. 10 mg Fe per ml.
E Iron Working Standard
Pipette 100 ml ironstock solution into
1 liter flask and dilute to mark with
distilled water.
1
ml
= 0.
010
mg
Fe
F Phenanthroline Solution
Dissolve 0.1 g 1, 10-phenanthroline mono-
hydrate, C12HgN2-H2O, in 100 ml distilled
water by stirring and heating to 80°C; do
not boil. Discard the solution if it darkens.
(Note that 1 ml of this reagent is sufficient
for no more than 0. 1 mg Fe).
G Working Solutions
1 Reducing reagent - one part NH2OH
added to two parts cone. HC1.
2 Color reagent - ten parts acetate
buffer added to four parts phenanthroline.
Note: The working solutions should be
made up daily.
II PROCEDURE
All glassware should be prerinsed with cone.
HC1.
A Take an aliquot of sample containing
0.01 - 0. 1 mg Fe. Dilute to 50 ml in
Erlenmeyer flask. Treat 0.0, 1.0, 2.0,
4. 0, 8.0, and 10 ml of iron working
standard in the same manner.
B Add 3 ml of reducing reagent. Boil on a
hot plate until volume reaches about 20 ml
(35 minutes).
C Cool. Add to 50 ml rough calibrated
Nessler tube containing 14 ml. color
reagent. Dilute to 50 ml. Mix thoroughly.
D After 15 minutes, read absorbance on
spectrophotometer using 1 cm cells at
510 m|j. and setting reagent blank at
0 absorbance.
E Plot concentration of Fe vs. absorbancy.
F Determine absorbance of unknown sample
and calculate concentration of sample
from standard curve.
*In Charge, Chemistry, Technical Advisory and Investigations Section, DWS&PC, and Chemist,
DWS&PC Training Activities, SEC. Reviewed December 1965.
CH. ME. FE. mn. la. 11. 64
20-8
-------
THE DETERMINATION OF MANGANESE BY PERIODATE OXIDATION
D. G. Ballinger and B. A. Punghorst*
I REAGENTS
A Sulfuric Acid, Concentrated
B Nitric Acid, Concentrated
C Phosphoric Acid, 85% Solution
D Potassium Metaperiodate, KIO.
E Silver Nitrate
F Manganous Standard
1 Dissolve 3. 2 g KMnO4 distilled water
and make up to a liter.
2 Heat solution for several hours near
the boiling point.
3 Filter through fritted glass filter.
4 Standardize against sodium oxalate.
G Manganous Working Solution
Calculate the volume of manganous stand-
ard (F) to dilute to 1 liter so that 1.0 ml
- 0.05 mg Mn. To this volume add
2-3 ml cone. H2SO4 + NaHSO3 until the
permanganate color disappears; boil to
remove excess SO2. Dilute to 1 liter.
II PROCEDURE
A Prepare manganese standards by pipetting
0.0, 1.0, 2.0, 4.0, 6.0 ml of manganous
working solution (I-G) into 125 ml
Erlenmeyer flasks. To the sample flask
add an aliquot of sample containing 0.05 mg
Mn - 0. 30 mg Mn.
B Add 5 ml concentrated H2SO4 and 5 ml
concentrated HNOg. Evaporate to SOg
fumes.
C Cool. Cautiously add 85 ml distilled
water. Cool again. Add 5 ml HNO3
and 5 ml H3PO4. Mix.
D Add 0. 3 g KIO4 and 20 mg AgNO3.
E Heat to boiling and keep at slightly below
boiling point for 10 min. or at least an
hour for small amounts of Mn.
F Cool and dilute to 100 ml in Nessler tubes.
Ill CALCULATIONS
A Prepare standard curve with mg Mn vs.
absorbancy at 525 m(j..
B Determine absorbance of unknown sample.
C Calculate mg Mn in unknown from standard
curve.
REFERENCE
1 Standard Methods for Examination of
Water and Wastewater. llth Ed.
APHA. AWWA. WPCF. 1960.
*In Charge, Chemistry, Technical Advisory and Investigations Section, DWS&PC, and Chemist,
DWS&PC Training Activities, SEC. Reviewed December 1965.
CH. ME. mm. 3. 11. 64
20-9
-------
PRINCIPLES OF EMISSION SPECTROSCOPY
J. F. Kopp* ^
I INTRODUCTION
The term "spectroscopy, " in its broadest
sense, refers to the study of the radiations
of the electromagnetic spectrum. As no
single instrument exists which will separate
radiation from all parts of the spectrum, it
is divided into regions which are related to
the different types of instruments capable of
producing or measuring waves of various
lengths. A diagram of the spectral dis-
tribution of energy is shown in Figure 1.
Emission spectroscopy is concerned with that
region wherein radiation can be sorted out
into a spectrum by means of prisms or grat-
ings, and includes the near infra-red, the
visible and the ultraviolet.
Emission spectroscopy has become an indis-
pensable part of the modern chemist's
analytical methods. With it he can analyze
a wide variety of substances both qualitatively
and quantitatively for trace elements regard-
less of valence states.
II THEORY
A Origin of Spectra - Excited atoms and
atomic ions emit light of definite wave-
lengths. The excitation of multiple
elements in a sample results in the simul-
taneous production of the spectra of all of
these elements.
During excitation by a thermal or electri-
cal source, the outer orbital electrons of
an element absorb energy and rise to
higher energy levels. These electrons
then return spontaneously to their normal
or ground states by a single jump or by a
series of jumps. The energy emitted
with each jump produces a spectral line
of characteristic wavelength and frequency
for the particular chemical element. The
combination of lines produced by the
excited atoms of the element thus provides
the emission spectrum of that element.
FIGURE 1
SPECTRAL DISTRIBUTION OF ENERGY
A R
x 10* 10* to4 ioz 10° 10-2 10-* i i 10-* lo-10 icr2 METERS
L i L 1 l A * l All j i . I. ll I i I i i
1
10' 3 5 7i '
— RADIO -j
11 13
1 INFRA-
r RED"*
I
ULTRA? "
VlOLEl
—ULTRA SONIC -
PERSEC
I S\ | GAMMA | COSMIC
HERTZIAN WAVES 1 / \ |*~ RAYS J RAYS~~
h-VISIBLE-}
|—X-RAYS—j
IINFIjAREDj VISIBLE j
WAVELENGTH X 1500 75060<)5004bo 300 250 200
REGION INCLUDED IN IMIIIIOMTIKCTROSCOPY
B
'.m^- METERS x 10"
10 ANGSTROMS
*Spectrochemist, Water Pollution Surveillance System, SEC. Reviewed December 1965.
CH.MET.es.1.5.65 21'1
-------
Principles of Emission Spectroscopy
B Types of Spectra
There are three types of emission spectra:
1 the line spectra produced by highly
excited atoms or atomic ions;
2 band spectra which originate with
highly excited molecules;
3 continuous spectra which result when
light is emitted by incadescent solids.
Ill INSTRUMENTATION
The emission spectrograph consists of
essentially four distinct functional parts,
the excitation source, the slit, the optical
system and the recording system. A diagram
of a Littrow type spectrograph is shown
in Figure 2.
A The production of line spectra for the
detection or the determination of the
elemental constituents in a sample, re-
quires an excitation source such as an
arc, a spark or a flame.
B The slit of a spectrograph permits only
a narrow beam of light of mixed wave-
lengths to enter the instrument.
C The optical system consists of a series of
lenses and either a prism or a grating, to
separate light rays of different wave-
lengths into the spectrum.
D The recording system can be either;
1 an eye piece as ie used in a spectro-
scope,
2 a photographic plate as is used in a
spectrograph, or
FIGURE 2
SCHEMATIC DIAGRAM OF A
LITTROW SPECTROGRAPH
EXCITATION
SOURCE
SLIT i
COLLIMATOR-
CAMERA LENS
MIRRORED
BACK
TOTAL
REFLECTING
PRISM
30-60-90*
QUARTZ PRISM
CAMERA
21-2
-------
Principles of Emission Spectroscopy
IV
a photoelectric cell as is used in
the direct reading spectrograph.
SELECTION OF EQUIPMENT AND
COST CONSIDERATIONS
A decision concerning the advisability of
installing a laboratory for spectrochemical
analysis, whether in an industrial plant,
research laboratory or state health depart-
ment might rest upon four main considerations:
A Whether the analytical problems encounter-
ed have no satisfactory solution apart from
spectroscopy,
B The time urgency of the work,
C The expected volume and diversity of
work,
D The cost of the initial investment, to-
gether with operating expenses.
V PROBLEMS AND TECHNIQUES
ASSOCIATED WITH EMISSION
SPECTROSCOPY
After the equipment has been set up properly
and is ready for operation there are many
parameters to be determined before the first
samples are analyzed.
A Source - arc or spark
There are several types of excitation
sources available as are shown in Table I.
Both the type of sample and the precision
and accuracy required for the analysis
will generally govern which type of
excitation method should be employed.
B Type of Electrode
The spectrographic electrode plays an
important part in the analysis. Some
important features are:
TABLE 1. EXCITATION SOURCES
TYPE
D. C. Arc
High Voltage
A.C. Spark
Low Voltage
A. C. Arc
Spark Ignited
Uni - Arc
High Voltage
A. C. Arc
PRECISION
Fair
Excellent
Good
Good
Fair
SENSITIVITY
Excellent
Fair
Good
Good
Excellent
RANGE
0.00001 -
1, 00%
0.01-30.0%
0.001 - 1.0%
0.001 - 1.0%
0.00001 -
1.0%
USE
Basic source for general
qualitative analysis - soils,
ores, oxides, slags, ashes,
etc. Highest sensitivity of
detection for trace elements.
Most stable. Use for alloy-
ing constituents in metals.
Solution techniques.
Quantitative determination
of residual impurities low
alloying constituents in
metals.
Combines precision of the
spark and the sensitivity of
the D. C. Arc.
Steadier than D. C, Arc -
Conductors and nonrefrac-
tory materials.
21-3
-------
Principles of Emission Spectroscopy
Density - the current and temperature
obtainable are dependent on the density
of the graphite.
Shape - the nature of the sample being
analyzed will usually determine the
shape and type of electrode.
Purity - trace analysis requires that
the electrodes be of very high purity.
C Exposure Conditions
Current, voltage, exposure time, etc.,
all play an important part.
D Type of Photographic Plate
The photographic emulsion most generally
used for quantitative analysis is the S. A.
#1. This emulsion responds to the
spectral range from 2400 to 4400 A. It
has high contrast, low background density
and low granularity.
The S. A. #2 emulsion is also desirable
for trace analysis when lower contrast,
higher speed and wider latitude are
needed. S.A.#2 also covers the range
from 2400 to 4400 A. Other photographic
emulsions, covering the wavelength range
from 2400 to 12000 A, are available, each
having specific features. Processing of
these photographic plates is extremely
important and directions of the manu-
facturer should be closely followed.
E Qualitative Analysis
In spectroscopy, qualitative analysis
is a relatively simple process although it
can be more time consuming than a
quantitative determination, especially if
a complete qualitative analysis is re-
quired. The presence or absence of over
60 of the chemical elements can be readily
determined by a simple inspection of the
resulting pattern of spectral lines. All
elements give specific lines when
sufficiently excited, which is the basis of
qualitative spectrochemical analysis.
F Quantitative Analysis
Quantitative spectrochemical analysis
is based on the fact that the amount of
light emitted by an element present at
very low concentrations is directly pro-
portional to the number of its excited
atoms present, if all other factors are
kept constant. The intensity of the
spectral line of the analysis element
(or degree of blackening of the photo-
graphed image of the line) in an unknown
sample is compared with the intensity
of the corresponding line in a standard
sample to provide an estimation of the
concentration of the element producing
that line.
G Internal Standard
In quantitative analysis the errors caused
by temperature fluctuations and wandering
of the arc are minimized by the use of the
internal standard technique. This consists
of comparing the intensity of a suitable
line of the analysis element in the standard
and in the unknown sample to a certain
selected line of another element whose
concentration is fixed in all samples.
As both the unknown and the internal
standard are a part of the same sample,
variations in the time of exposure, plate
characteristics and developing conditions
will not affect the relative density of the
two lines which are equal in intensity in
the light source.
Working curves are established by plotting
the intensity ratio of the analysis line to
the internal standard line versus the con-
centration of the analysis element.
VI APPLICATION OF EMISSION SPECTRO-
SCOPY TO WATER ANALYSES
Trace elements in water originate from a
variety of sources but can be classified into
three principal groups:
A Elements contributed by soluble materials
chemically weathered from soil and rocks.
21-4
-------
Principles of Emission Spectroscopy
B Elements that are selectively concentrated
by vegetation and find their way to surface
waters following decay and run-off.
C Industrial sources, especially those de-
voted to mining, alloying, and cleaning and
plating of metals.
These sources may contribute significant
quantities of trace elements to surface
waters both in populated and unpopulated
areas. The detection and measurement
of these trace elements is difficult with
conventional analytical procedures because
they are not adapted to large numbers of
samples and in some cases are not sensi-
tive enough. The use of a spectrographic
procedure, however, for routine moni-
toring of raw waters is admirably suited
to the purpose since a large number of
elements may be determined simultaneously
with excellent accuracy. Table 2 lists
those elements routinely looked for and
their concentration ranges.
TABLE 2. WAVELENGTHS AND CONCENTRATION RANGES OF
ELEMENTS ADDED TO SYNTHETIC WATER MATRIX MATERIAL
Element
Cadmium
Barium
Beryllium
Lead (high)
Lead (low)
Chromium
Tin
Antimony
Manganese
Iron
Nickel
Bismuth
Molybdenum
Vanadium
Copper
Zinc
Cobalt
Silver
Wavelength, A
2280.0
2335.3
2348.6
2663.2
2833. 1
2677.2
2840.0
2877.9
2933. 1
2973.2
3003.6
3067.7
3170.3
3183.4
3274.0
3345.0
3453.5
3280.7
Micrograms per
Electrode Con-
centration Range
0.03 - 1.0
0.02 - 10.0
0.0005 - 0. 1
0.05 - 10.
0.05 - 1.0
0.01 - 1.0
0.02 - 1.0
0.1 - 1.0
0.04 - 1.0
0.02 - 1.0
0.02 - 1.0
0.05 - 0.1
0.02 - 0.1
0.02 - 1.0
0.003 - 1.0
5.0 - 10.0
0.02 - 1.0
0.0005 - 0.05
21-5
-------
Principles of Emission Spectroscopy
Trace elements, whether in raw or
finished water, are generally present in
concentrations too low to measure directly
with the spectrograph. A means of con-
centration, therefore, is necessary before
the examination can be completed. This
can be accomplished in several ways, i.e.,
evaporation, precipitation, ion exchange,
etc.
VII PREPARATION OF STANDARDS
As the major constitutents of a sample are
important factors that affect the burning
qualities of the arc and the intensities of
resulting spectral lines, it is important that
the composition of the standard matrix ma-
terial approximate the samples as closely
as possible. This is accomplished by pre-
paring a synthetic matrix to approximate the
average composition of waters of North
America as given by F. W. Clarke (see
references) and consists of 20% calcium, 5%
magnesium, 7. 5% sodium and 2% potassium.
The nitrates of these elements are combined
in a single solution using double distilled
water and to this synthetic matrix are added
increasing amounts of the analysis elements
over the concentration range.
VIII ACCURACY
The average error in quantitative emission
spectroscopy using the photographic process
is generally between + 5 - 10%. In the direct
reading process, however, where photo-
multiplier tubes are substituted for the
photographic plate, an average error of
+ 2% is possible. This accuracy is more
than satisfactory considering that determi-
nations of trace elements are being made in
the parts per billion range.
A Water Pollution Surveillance System Results
The spectrographic measurement of
trace metals in surface waters of the
United States has been performed routinely
by the System since 1958. Approximate
levels of detection for various water
systems are shown in Table 3.
Of the 17 elements included in the spectro-
graphic examination of those major water
systems shown (during the years 1958 -
1962), antimony, beryllium, bismuth,
cadmium, cobalt, tin and zinc were
either detected infrequently or not at all.
The frequency of detection for other
elements is shown in Table 4.
It should be emphasized that in quantitative
spectrographic analysis the best possible
standards must always be prepared. A good
spectrographer is only as good as his
analyses and his analyses are only as good
as his standards. Spectrographic standard
solutions are prepared from Reagent Grade
chemicals. The oxides are preferred where
possible; however, the nitrates or chlorides
will usually suffice. Occasionally, it is
necessary to use special spectroscopically
pure grades of certain metals. Nitric and
hydrochloric acids should be re-distilled,
and double distilled water is also suggested
where possible. Borosilicate glassware,
cleaned with both chromic and nitric acids,
is used throughout.
B Analytical Reference Service Results
A sample containing eight trace metals at
concentrations shown in Table 5 was
analyzed by sixty-six participating agencies
using a number of analytical methods. The
mean results of all sixty-six laboratories
in addition to those laboratories employing
spectrographic procedures are shown in
Table V. It is apparent from this summary
that the spectrograph can be used to
splendid advantage on water samples.
21-6
-------
Principles of Emission Spectroscopy
TABLE 3. APPROXIMATE LEVELS OF DETECTION
ACHIEVED FOR VARIOUS WATER SYSTEMS
(tig/D
Group
I Ag, Be
II Ba, Cr,
Cu, Fe,
Mn, Mo,
Ni, Pb, V
Cd, Co, Sn
m Bi, Sb
IV Zn
Colorado
0.3-0.5
2-20
30 - 60
2500 - 3500
Columbia
O.Q3 - 0.05
1-5
5-10
300 - 500
Great Lakes
0.05 - 0. 1
1-5
5-10
400 - 600
Ohio
0.05 - 0. 1
1 - 10
10 - 20
600 - 800
Mississippi
0. 1 - .2
2-20
20 - 30
800 - 1000
Missouri
0. 1 - 0.2
1 - 10
20 - 40
1500 - 2500
TABLE 4. FREQUENCY OF DETECTION OF TRACE ELEMENTS
IN CERTAIN WATER SYSTEMS
(1958 - 1962)
SYSTEM
Colorado River
Columbia River
Great Lakes
Mississippi River
Missouri River
Ohio River
Average
ELEMENT %
Ba
79
94
95
100
96
94
93
Cr
12
87
32
23
10
20
31
Cu
61
100
98
96
85
91
89
Fe
82
100
98
95
92
100
95
Mn
0
13
4
11
6
17
9
Mo
82
90
50
68
65
37
65
Ni
9
48
39
23
8
31
26
Pb
6
29
20
11
3
11
13
V
9
3
0
0
2
0
2
Ag
0
0
14
12
57
60
24
21-7
-------
Principles of Emission Spectroscopy
TABLE 5. SUMMARY OF RESULTS OBTAINED BY SPECTROGRAPHIC
PROCEDURES ON ARS SAMPLE; WATER METALS, NO. 2
Element
Al
Cr
Cu
Fe
Mn
Cd
Zn
Pb
Amount Added
mg/1
1.80
0. 18
0.42
0.62**
0.25
0.24
0.90
0. 18
Mean of
66 Labs
2.21
0. 14
0.43
0.44
0.28
0.26
0.94
0.20
AMOUNT RECOVERED mg/1
Lab #71126
1.8
0. 18
0.29
0.46
0.25
0.25*
0.68
0.27
NWQN Lab
2.30
0. 13
0.38
0.40
0.23
0.30
0.94
0. 19
Lab #1615
3. 10
0. 17
0.41
0.75
0.38
0.50
0.88
-
This result was erroneously included under Lab #7112 A in the ARS report.
**
Iron value was adjusted to mean value as suggested on page 47 of ARS report.
SUMMARY
The theory and instrumentation of emission
spectroscopy are presented. Consideration
is given to the problems and techniques
associated with trace element analysis in
water using the emission spectrograph.
REFERENCES
Emission Spectroscopy
1 Nachtrieb, Norman H. "Principles and
Practice of Spectrochemical Analysis. "
McGraw-Hill Book Co., Inc. 1950.
2 Clark, George L. "The Encyclopedia of
Spectroscopy. " Reinhold Publishing
Corp. 1960.
Water
Kroner, Robert C., and Kopp, John F.
"The Occurrence and Significance of
Trace Elements in Surface Waters. "
Presented at the Meeting of the Iowa
Section. American Water Works
Association, Sioux City, Iowa. Oct.
16-18, 1963.
Kopp, John F., and Kroner, Robert C.
"A Direct Reading Spectrochemical
Procedure for the Measurement of
Nineteen Minor Elements in Natural
Water. " In Press.
"National Water Quality Network, Annual
Computation of Data. " 1960-1961. U. S.
Dept. of Health, Education and Welfare,
Public Health Service Publication,
No. 663.
21-8
-------
Principles of Emission Spectroscopy
Haffty, Joseph "Residue Method for
Common Minor Elements. " U.S.
Dept. of Interior, Geological Survey,
Water Supply Paper, No. 1540 A. 1960.
Silvey, W. D., and Brennan R. "Concen-
tration Method for Spectrochemical
Determination of Seventeen Minor
Elements in Natural Water. " Anal.
Chem., 34, 784. 1962.
Skougstad, M.W., and Horr, C.A.
Occurrence and Distribution of
Strontium in Natural Water. " U.S.
Geological Survey, Water Supply Paper
No. 1496 C. 1963.
Clarke, F. W. "The Composition of the
River and Lake Waters of the United
States." U.S. Geological Survey,
Papers No. 135. 1924.
21-9
-------
THE POLAROGRAPHIC DETERMINATION OF COPPER,
CADMIUM, NICKEL, AND ZINC
C. E. Stephan*
I SOURCES AND SIGNIFICANCE
A Sources
(1)
All are produced in mining and refining of
ore and manufacture of the metals and
alloys. All are found in metal plating
wastes. In addition, copper salts are
used as algicides.
B Significance
All can interfere to some extent with the
efficient operation of sewage treatment
plants. * All can produce undesirable
taste in drinking water, and cadmium is
very toxic to humans. Individually
copper, cadmium, and zinc are toxic to
many forms of aquatic life at a concentra-
tion of 1 mg/liter (1 ppm), whereas
nickel can often be tolerated at the 10 mg/
liter level. But there have been many
reports that combinations of the metals
are more toxic.
II FACTORS INFLUENCING THE CHOICE
OF AN ANALYTICAL METHOD*5' 6» 7)
A Sensitivity
B Accuracy and Reproducibility
C Expense of Equipment and Reagents
D Time Requirements
E Sensitivity to Interferences and Ease of
Removal of Interferences
F Specificity
G Simultaneous Determination of Several
Substances
III SAMPLE PREPARATION
A Total, Dissolved, or Reactive Metal
B Removal of Interferences
1 Organic
rfJJ >
a Wet ashing(8) r*~'">i
b Dry ashing ;, ' - '~''f "
1) High temp. <9> 10> n>
2) Low temp. *12)
2 Inorganic
a Solvent extraction
b Complexation
c Electrolysis
d Ion-exchange
IV CHOICE OF ELECTROLYTE*13' 14)
A Well-Formed Waves
B Separation of Waves
C Removal of Oxygen
D Electrolytes Used
(15)
1 Ballinger-Hartlage
2. ml cone. HC1
10. ml distilled H2O
8. ml cone. NH4OH
0.5 ml 0. 2% Triton X-100
20. 5 ml + 0. 5 g Na2SO3
71'
*Chemist, Aquatic Biology Section, Basic and Applied Sciences Branch, DWSPC, SEC.
CH. ME. lla. 12. 65 22-1
-------
polarographic Determination
(16)
2 Mount
50 ml cone. HC1
200 ml cone. NH4OH
40 ml 0. 04% Triton X-100
210 ml distilled H2O
500 ml + Na2SO3
3 Sirois(17)
1. 0 M. NH4OH
1.0 M. NH4C1
0. 1 M. Na2SO3
0. 01 % Gelatin in H2O (""-*
4 Stephan
(18)
10 ml cone. HC1
40 ml cone. NH4OH
g Na2SO3
. 05 g Gelatin
10
450 ml H2O
500
ml
^
V ANALYTICAL PROCEDURES
(19)
A Sewage Samples for Zinc
1 Weigh sample into 50 ml beaker and
evaporate to dryness.
2 Add 5 ml HNOg and 2 ml H2SO4 and
boil to dryness (Use HC1O4 on resis-
tant samples).
3 Add electrolyte no. 2. Filter.
4 Run polarogram.
B Tissue Samples for Cadmium and
Zinc<16'W
1 Weigh tissue in crucible.
2 Ash for 4 hours at-£oO°C.
3 Dissolve ash in acid.
4 Separate cadmium and zinc on ion
exchange columns.
5 Evaporate to dryness.
6 Add electrolyte no. 2 and run *
polarogram.
(17 21
C Plant samples for Copper and Zinc '
1 Weigh sample in beaker
2 Ash at 500°C for 4 hours.
3 Add hydrochloric acid and boil to
dryness.
4 Add electrolyte no. 3. Filter.
5 Run polarogram.
D Water Samples for Copper, Cadmium,
Nickel, and Zinc* '
1 Place aliquot in beaker and acidify.
2 Evaporate to dryness.
3 Ash, add HC1, and boil to dryness,
if necessary.
4 Add electrolyte no. 4.
5 Transfer to 10 ml beaker.
6 Run polarogram.
REFERENCES
1 Kroner, R. C. and Kopp, J. F. JAWWA
57: 150. 1965.
2 Earth, E. F. et al. JWPCF 37: 86. 196:
3 PHS Drinking Water Standards. PHS
Publication No. 956. 1962.
4 Water Quality Criteria. Publication No.
3-A, Resources Agency of California.
1963.
5 Margerum, D. W. and Santacana, F.
Anal. Chem. 32: 1157. 1960.
6 Kroner, R. C. et al. JAWWA 52: 117.
1960.
7 O'Connor, J. T. and Renn, C.E. JAWW/
55:631. 1963.
22-2
-------
Polarographic Determination
8 Smith, G.F. Talanta 11: 633. 1964.
9 Pijck, J. et al. Internal. J. Appl.
Radiation Isotopes 10: 149. 1961.
10 Alexander, G. V. Anal. Chem. 34: 951.
1962.
11 Gorsuch, T.T. Analyst 87: 112. 1962.
12 Low Temperature Dry Asher, Tracerlab,
2030 Wright Avenue, Richmond, Calif.
13 Bush, E.L. and Workman, E.J. Analyst
90: 346. 1965.
14 DuBois, L. and Monkman, J. L. Am.
Ind. Hygiene Assoc. J. 25: 485. 1964.
15 Ballinger, D. G. and Hartlage, T.A.
Water and Sewage Works. 109: 338.
1962.
16 Mount, D. L Trans. Amer. Fisheries
Soc. 93: 174. 1964.
17 Sirois, J. C. Analyst. 87: 900. 1962.
18 Stephan, C.E. Unpublished.
19 McDermott, G. N. et al. Ind. Waste
Conf. Purdue Univ. Ext. Ser. 112:
461. 1962.
20 Mount, D.I. and Stephan, C. E. Unpublished.
21 Robertson, G. Analyst. 89: 368. 1964.
-------
THE DETERMINATION OF PHENOLS
J. W. Mandia*
I DEFINITION AND SIGNIFICANCE
A Definition
The phenolic compounds in water chemistry
collectively referred to as phenols are
defined as those hydroxy derivatives of
benzene, or its condensed nuclei, which
can be determined colorimetrically by the
aminoantipyrine or Gibbs method.
All chlorination products may contribute
to the intensity of taste and odor.
At maximum taste and odor intensity
the major contributor is a hitherto
overlooked compound 2, 6-dichlorophenol.
The chlorine-to-phenol ratio at maximum
intensity of taste and odor is 2:1. The
proportion of 2, 6-DCP was greatest at
the 2:1 chlorine-to-phenol ratio.
B Significance
Phenol and chlorinated derivatives in
water affect fish and water quality.
1 Fish
The threshold limit of toxicity at
infinite time for certain species of
fish is of the order of a few milligrams
per liter. Some chlorinated phenols
exhibit toxicity in concentrations as
low as 0. 2 mg/1.
2 Fish flesh tainting
Fish living in waters of lesser
phenolic concentration can acquire
an unpleasant and obnoxious taste.
Ill STANDARDIZATION OF PHENOL -
Bromate-bromide Method
A Reaction
6HC1
2KBrO3 + 2KBr ^ 2Br2 + 4KC1 + 3H2O
OH
3Br
u +
3HBr
Br
B Calculations
Equivalent wt of phenol =
3 Water quality
The presence of as little as 1 ng/1 of
the chlorinated phenols can impart a
taste to drinking water.
atomic wt of phenol
atoms of Br /molecule of unreacted phenol
£1
94. 11
= 15.685 eq. wt.
II CHLORINE DERIVATIVES OF PHENOL
CAUSING TASTE AND ODOR
A Progressive Chlorination of Phenols
2- and 4- CP: 2,4 dichlorophenol,
2, 4, 6 trichlorophenol and 4, 4
dichloroquinone, 2, 6 dichlorophenol
1 ml of 0. IN bromate-bromide = 0.1
millequivalent (meq)
Meq X' equivalent wt = milligrams (mg)
0. IN bromate-bromide = 0.1 X 15.685
or 1. 5685 mg of phenol
*Chemist, DWS&PC Training Activities, SEC. Reviewed December 1965.
CH.PHEN. 32a. 12. 63 23-1
-------
The Determination of Phenols
1 ml of 0. IN bromate -bromide = 4 ml of
0. 025N thiosulfate;
1 ml of 0.025N thiosulfate = 1.5685/4
= 0. 3923 mg of phenol
(AB-C) = amount of brominating agent
consumed by the sample,
expressed as ml of 0. 0 25N
thiosulfate.
A = ml of 0. 025N thiosulfate for
each 10 ml portion of bromate -
bromide reagent, used for the
blank.
B = No. of 10 ml portions of the
bromate-bromide reagent used
for the blank.
C = ml of 0. 025N thiosulfate used
to titrate the excess brominating
reagent in the sample.
For a determination based on 50 ml of
sample, mg phenol/50 ml of sample
= (AB - 0(0. 3923)
impurities. The rate of volatilization of «
the phenols is gradual, so that the volume
of the distillate must equal that of the sample
being distilled.
A Reagents
1 Copper sulfate
CuSC>4 forms CuS thus preventing the
formation of H^S which interferes with
the determination. It also prevents
biochemical degradation.
2 Phosphoric acid
Acidification of the sample with
H3PO4 assures the presence of the
copper ion.
B Interferences
Oxidizing agents as detected by the starch-
iodide test, are removed immediately
after sampling by the addition of an excess
of ferrous sulfate or sodium arsenite.
mg Phen01 = (AB- 0(0. 3923) (1°°0)
liter of sample
50
V THE 4-AMINO ANTIPYRINE METHOD
= (AB - C)(7.846)
A Procedure
IV PHENOL DISTILLATION
Phenols are distilled at more or less
constant rate from the nonvolatile
Purified phenols but not paracresol react
with 4-amino antipyrine at a pH of 10.0
in the presence of ferricyanide to form
an antipyrine dye. The dye is extracted
from aqueous solution with chloroform
and the absorbance read at 460 m|J..
Reaction
CH3-N
CH3-C = C-NH2
pH - 10
K-Fe (CN)
3 6
OH
CHQ-N C=0
o
CH -C = C-N
O
RED
= O
23-2
-------
The Determination of Phenols
The reagent 4-aminoantipyrine reacts
with phenols in the presence of an
oxidizing agent to form a colored
compound sensitive to a few ppb.
With all of the simple phenols the
reaction product is the same color
(red) although the response with
different phenols varies. Phenol,
CgHgOH is most sensitive and all
results are customarily reported in
terms of CgHgOH.
The reaction product is soluble in
chlorinated hydrocarbons. Chloroform
is the solvent of choice by reason of
extraction efficiency, stability, cost,
toxicity, etc.
B The Variables in the 4 AAP, Amino
Antipyrine Method
1 The reaction is carried out at pH 10
to minimize interference from aniline
and other aromatic amines.
2 Time of color formation is not a
variable since the reaction takes place
immediately.
3 The reagents, 4-aminoantipyrine and
the oxidant, potassium ferricyanide,
are fairly stable. Refrigeration in the
absence of light prolongs reagent life
by a matter of weeks. Variations in
batches of 4 AAP have been observed
but are not troublesome.
4 The reaction product fades slowly over
a period of hours in the aqueous phase.
The extracted material fades very
slowly but the blanks tend to darken.
The effect is not noticeable for a period
of several hours.
Because of the stability of the reagents
and the uniformity of color response it
is possible to prepare a semi-permanent
standard curve.
1 For phenol concentration under
1.0 mg/liter all samples are extracted
with chloroform to increase sensitivity.
For phenol concentrations above
1. 0 mg/liter readings may be made
in the aqueous phase. A blank and one
standard at 5. 0 mg/liter may be run
as a check on a previously prepared
standard curve.
Using the chloroform extraction
procedure the response is linear
from zero up to about 1.0 mg/liter.
For aqueous phase readings the
response is linear from about
0. 1 mg/liter of phenol.
D Substituted Phenolics
The 4-aminoantipyrine reaction is
applicable to phenolic compounds in which
the para-position is not blocked by an aryl,
alkyl, nitro, benzoyl, nitroso or carbonyl
group. These do not give color when
present in para-position.
1 The color produced is of varying
intensity. Phenol, itself gives the
greatest density. Consequently, when
using a standard curve referred to
phenol itself, a mixture of substituted
phenolics will give a minimum density
value.
2 Density of substituted phenolics
compared to phenol:
Compound
Phenol
O-Cresol
M-Cresol
P-Cresol
Density Compared to
Phenol
100
74
69
3
REFERENCES
Aminoantipyrine Procedure
1 Emerson, E. , Beachham, H.H., and
Beesle, L.C. The Condensation of
Aminoantipyrine. II A New Color Test
for Phenol. J. Org. Chem. 8:417.
1943.
23-3
-------
The Determination of Phenols
Ettinger, M. B. , Ruchhoft, C.C., and 4 Mohler, E.F., Jacor, L.N. Comparison •
Lishka, R. J. Sensitive 4-Amino- of Analytical Methods for Determination
antipyrine Method for Phenolic of Phenolic Type Compounds in Water
Compounds. Anal. Chem. j!3_:1783. and Industrial Wastes Water.
1951. Presented at 130th Meeting of ACS.
Atlantic City. September 16-21, 1956.
Dannis, M. Determination of Phenols
by the 4-Aminoantipyrine Method.
Sew. and Ind. Wastes. 23:1516. 1951.
23-4
-------
LABORATORY EXERCISE - ANALYSIS OF WASTE SAMPLE FOR PHENOL
R. C. Kroner*
I PRELIMINARY DISTILLATION
A Reagents
1 Phosphoric Acid, 10%
2 Copper Sulfate, 10%
B Procedure
1 Place 250 ml of sample in a beaker and
add 1 ml phosphoric acid solution.
Using test paper, check to see that
pH is <4.
2 Add 1 ml of copper sulfate solution and
transfer to the distilling flask. Con-
nect flask to condenser and start dis-
tillation. Collect distillate in anErlen-
meyer flask marked at 250 ml.
3 When approximately 200 ml of the dis-
tillate has been collected, withdraw the
heat and add 50 ml distilled water to
the distilling flask. Continue distilla-
tion until 250 ml has been collected.
Swirl the distillate in the flask to mix.
II DETERMINATION OF PHENOL BY 4-
AMINOANTIPYRINE METHOD
A Reagents
1 4-Aminoantipyrine, 2% aqueous
U
2 Potassium ferricyanide solution, 8%
aqueous.
3 Ammonium chloride buffer, pH 10 -
dissolve 67. 5 g NH4C1 in 570 ml cone.
NH4OH and dilute to 1 liter with dis-
tilled water.
4 Standard phenol solution, 1 ml = 1 mg
B Procedure
1 Prepare a 5 mg/1 phenol standard by
diluting exactly 5 ml of the standard
phenol solution to 1 liter with distilled
water.
2 Measure duplicate 100 ml portions of
the 5 mg/1 standard into labelled Erlen-
meyer flasks.
3 Prepare a blank by measuring 100 ml
distilled water into a labelled Erlen-
meyer flask.
4 Measure 100 ml of the distillate into
a labelled Erlenmeyer flask.
5 To the standards, blank, and sample add:
2 ml ammonium chloride buffer
2 ml 4-AAP reagent
2 ml potassium ferricyanide reagent
Mix well after each addition
6 Measure the optical density of each of
the standards, blank, and sample at
510 m^, using a 1. 0 cm cell.
7 Record the readings on the data sheet
and compute the concentration of phenol
in the original sample, in mg/1.
Ill DETERMINATION OF PHENOL BY 4-
AMINOANTIPYRINE METHOD (EXTRACTED)
A Reagents
1 4-Aminoantipyrine solution, 2% aqueous
2 Potassium ferricyanide solution, 8%
aqueous
3 Ammonium chloride buffer, pH 10 -
dissolve 67. 5 g NH4C1 in 570 ml cone.
NH4OH and dilute to 1 liter with dis-
tilled water.
4 Standard phenol solution, 1 ml = 1 mg
*In Charge, General Laboratory Services, Water Quality Section, DWS&PC, SEC. Reviewed
December 1965.
CH.PHEN. 31. 11. 61
23-5
-------
Laboratory Exercise - Analysis of Waste Sample for Phenol
5 Chloroform, Reagent grade.
B Procedure
1 Prepare a 0. 05 mg/1 (50 ppb) phenol
standard by diluting exactly 5.0 ml
of the standard phenol solution to 1 liter
with distilled water, then diluting 10 ml
of this solution to 1 liter. The final
standard contains 0. 05 mg/1.
2 Transfer 500 ml of the above standard
solution to a 1 liter separatory funnel.
3 Prepare a blank by measuring 500 ml
distilled water into a 1 liter separatory
funnel.
4 Dilute 10 ml of the distillate to 1 liter
with distilled water, Transfer 500 ml
of the diluted sample to a 1 liter separa-
tory funnel.
5 To the standard, blank, and sample in
the separatory funnels add:
3 ml ammonium chloride buffer
3 ml 4-AAP reagent •
3 ml potassium ferricyanide reagent
Mix well after each addition
Extract the solutions serially with 15,
10, and 5 ml volumes of chloroform.
Collect the chloroform extract by
filtering through paper into a 25 ml
graduate. Make up to 25 ml with
chloroform.
Measure the optical density of the stand-
ard, blank, and sample at 460 mji, using
a 5. 0 cm cell.
Record the readings on the data sheet
and compute the concentration of phenol
in the original sample by multiplying the
measured concentration by 100.
23-6
-------
LABORATORY EXERCISE - ANALYSIS OF WASTE SAMPLE FOR PHENOL
DATA SHEET
4-AMINOANTIPYRINE METHOD (AQUEOUS)
Flask
Blank
Standard
Standard
Sample
O. D.
4-AMINOANTIPYRINE METHOD (EXTRACTED)
Funnel
Blank
Standard
Sample
O. D.
Cone, phenol, mg/1
0
5.0
5.0
Cone. phenol, ppb
0
50
Cone, phenol in original sample = ppb in diluted sample X 100.
23-7
-------
SOURCES, EFFECTS AND ANALYSIS OF CYANIDES
F. J. Ludzack*
I SOURCES EFFECTS AND ANALYSIS OF
CYANIDES
A Cyanides commonly are a first considera-
tion in fish kills just as phenols are com-
monly credited for objectionable water
taste or odor. The fact that many other
pollutants or conditions can and do cause
similar effects may not prevent unjust
accusation. Care must be used in dis-
charge and records to protect the public
and the processor.
B Fish toxicity has been reported at 0.025
mg/1 CN. Microorganisms generally are
more tolerant. Certain organisms are
adaptable to the destruction of cyanides.
Activated sludge or trickling filters suc-
cessfully destroyed 60 mg/1 feed cyanide
producing a nitrified effluent with negli-
gible cyanide. Cyanide is an enzyme
blocking agent, therefore, toxicity depends
upon reactivity of essential enzymes and
cyanide and the relative fraction of in-
activated enzyme. HCN is similar in
action to CO in human toxicity.
II INDUSTRIAL SOURCES OF CYANIDES
A Metal plating
B Case hardening
C Metal cleaning baths
D Silver and gold refining
E Gas scrubbers from pyrolytic processes,
(coking, refining, blastfurnaces.)
F Rubber, acrylic fiber, plastic industry.
Mainly as copolymers with nitriles.
G Chemical process intermediates
III CYANIDE TREATMENT
A Lagooning - may lead to complexation,
oxidation, polymerization or volatilization.
B Acidification and aeration transfers HCN
to the air.
C Lime-sulfur treatment forms less toxic
SCN".
D Complexation with heavy metals - of
doubtful value.
E Precipitation as heavy metal salts - re-
quires close control.
F Oxidation with KMnO4, ClOg, O3, C12.
Alkaline chlorination currently gives best
results. Cr oxidation conditions have
not been found. '
G Biochemical oxidation for low
concentrations.
v
H Ion exchange
I Electrolytic oxidation for high
cone ent rations.
IV Reactivity and the variety in form of cya-
nides cause major difficulties in treatment or
analysis and require individual handling of
each problem.
A Major forms of cyanide include:
1 Simple soluble cyanides -
H+Na+K+NH4+
2 Simple insoluble cyanides -
Ni(CN)2 Zn, Cd, etc.
3 Complexed cyanides - association
products of 1 & 2.
*Chemist, Chemistry and Physics Section, Basic and Applied Sciences Branch, DWS&PC, SEC.
CH.Cy.18a. 12. 65 24-1
-------
Sources, Effects and AnalysiSLof Cyanides
4 Organic cyanides - R-CN (Nitriles).
5 Cyanates (OCN~) and thiocyanates
(SCN~).
6 Organic compounds degrading to form
HCN during analysis (glycine).
B Item Al is the only one that can be treated
or analyzed efficiently. Other forms
must be converted to simple soluble cya-
nides during or prior to treatment or
analyses. Other forms of cyanide may be
considered as interferences.
V ANALYTICAL INTERFERENCE CONTROL
A Insoluble and complexed cyanides gener-
ally can be converted to simple cyanides
by the Serfass distillation technique in the
presence of HgC^, MgCl-2 and acid. Cer-
tain copper and cobalt complexes and prob-
ably others are not converted completely
to simple cyanides except after extended
distillation periods.
B Sulfides - precipitate with PbCO3 before
analysis.
C Fatty acids or other organic turbidity -
frequently may be extracted with iso-
octane, hexane or chloroform from slightly
acid solution.
D Amines - extract from slightly alkaline
solution as in V, C. The three solvents
will not extract detectable HCN.
E Oxidizing agents may destroy HCN. Test
with starch-Kl paper and titrate with
sodium sulfite to a negative test before
analysis.
F Turbidity - remove by acid distillation.
Trace quantities may be controlled by
extraction of the cyanide color complex.
G Color-try extraction, distillation or both.
H SCN" and OCN" acid distillation destroys
them.
Organic cyanides or organics degrading to
cyanide under analytical conditions gen-
erally interfere to a degree depending
upon the individual constituent. No general
method of removal can be recommended.
Results are open to question as these con-
stituents may produce false cyanide tests.
Nitriles and organic nitrogen compounds
may produce low level interference. Acid
distillation may help.
Buffer capacity of the sample may raise
the pH above 8. 0 during color formation
which decreases resulting absorbance'
materially.
VI ESTIMATION OF CYANIDE
A Silver nitrate titration using p-dimethyl
aminobenzalrhodanine indicator is recom-
mended when cyanide is 1 mg/1 or higher.
B Pyridine pyrazolone color test recom-
mended when CN is less than 1 mg/1.
May use aqueous or extracted absorbance
reading. Indicated CN of approximately
0.005 mg/1 is questionable.
VII GENERAL CONSIDERATIONS
A Available methods of analysis cannot sep-
arate so called "free" and "total" CN.
Certain complexes are stable enough to
prevent complete analytical recovery.
Part of the complexed cyanide invariably
is included with the simple cyanides.
Easily hydrolyzed cyanides will be recov-
ered in high yield; stable complexes may
not.
B Sample preservation is never 100% effec-
tive. Complexes may form or dissociate
in storage. HCN may volatilize. Prompt
analysis is recommended. Alkali addition
to pH 11 or above and cold storage is the
next best approach.
C Importance of distillation cannot be over
estimated. It should not be omitted unless
check tests with and without distillation
are comparable.
24-2
-------
Sources, Effects and Analysis of Cyanides
D Serfass distillation with HgCl2 and MgCl2
is recommended. One hour of reflux will
result in recovery of easily hydrolyzed
cyanides in high yield along with part of
the stable complexed cyanide. If cyanide
appears during a second one hour reflux
stable cyanides are indicated and low re-
covery may be expected.
E Results should be expressed in terms of
the CN ion even though it generally does
not exist in the sample as such except in
small proportions.
/Ill COMMENTS, CN PROCEDURES, 11TH
ED. OF STANDARD METHODS
A Comments on "free" cyanide may be
misleading.
i II
B Certain complexes containing Cu Cu
mixtures may be highly resistant to acid
distillation although most copper cyanides
are recovered in good yield.
C UndissociatedHCN is generally considered
more toxic to fish than the CN" ion. Rel-
atively little CN" occurs in samples except
at high pH values.
D Use of the work "screening" in place of
"interference control" is popular but
somewhat confusing.
E The color test is not particularly sensitive
to "salt" concentration unless the salt
buffers the color mixture to a pH greater
than 8.0. Color will not form consistently
in strongly alkaline solutions. See item N.
F Kruse and Mellon also suggested extraction
of cyanide from aqueous samples. The
best partition coefficient found was about
0.4 for transfer of HCN to the organic
solvent which does not contribute to a high
CN yield. Removal of interference as
described is effective.
G Apparatus item 2. 1 may be obtained as
Sargent No. 34087 including the air inlet
(item 2. 2). The Fisher Milligan gas
washer is difficult to seal, drain, and
rinse, and is not recommended. Sargent
No. 39623 is the one indicated for item 2. 4.
H Reagent 3. 2 is likely to be hard to pre-
pare as it is very close to a saturated
solution at room temperature. The
strength of mercuric chloride is not
critical - from 3 to 5 per cent solution is
adequate and easier to dissolve.
I It is not advisable to increase the air
rate if the liquid starts to back up into
the air inlet. Decrease the heat instead.
A high air rate is likely to blow cyanide
through the absorber. An air rate below
that of bubble coalescence in the spiral
is suitable. A heating mantle with a
powerstat control may be set to avoid
difficulties during warmup.
J 5 ml of acid is adequate for up to 1000 ml
of sample providing excess alkali has been
neutralized. The suggested 5 ml/100 ml
solution may result in low yields. Distil-
lation pH of about 1.5 is desirable.
K Item 1. 2 titration method. Distillation
will not remove interference such as H^S,
fatty acids, or volatile color.
L Item 3.5 Colorimetric Method. If the
chemical does not dissolve immediately
the analyst can expect difficulties due to
changes in storage. Replace with new
chemical.
M Item 3. 8 Use Eastman 6969, 3, 3'-dimethyl-
1, 1'diphenyl (4, 4'-bi-2-pyrazalene)-5,
5' dione.
N Color Procedure item 4.1. Dilute with
distilled water. If the suggested NaOH
solution is used here it will raise the pH
above the point of maximum color
development.
REFERENCE
Standard Methods, Water and Wastewater,
4/J' Hth Edition includes a good reference
list. Additional biological treatment
information may be found in the following:
NITRILES
1 Sew. & Ind. Wastes 31, No. 1, 33, 1959.
24-3
-------
Sources, Effects and Analysis of Cyanides
2 Proc 13 Ind. Waste Conf., Purdue Univ., 4 J.W.P.C.F. 33, p. 492, May, 1961.
Ext. series 96, p. 297, May, 1958. Act Sludge tr. of Cn, OCN, and SCN.
3 Proc. 14th Ind. Waste Conf., Purdue 5 Proc. 15th Ind. W. Conf. Purdue Univ.
Univ., Ext. series 104, p. 547, May, Eng. Ext. series 106, p. 439, May,
1959. 1960.
24-4
-------
LABORATORY PROCEDURE FOR CYANIDE
B. A. Punghorst*
Serfass Distillation Procedure
I REAGENTS
A Sodium hydroxide, 4% aqueous solution.
B Mercuric chloride, 5% aqueous solution.
C Magnesium chloride solution - add 51 gms
of MgCl2- 6H2O to flask, dissolve in
minimum quantity of water and make up
to 100 ml volume.
D Sulfuric acid, concentrated.
H Drain the contents of the gas scrubber
into a 250 ml volumetric flask and bring
to the mark with washings from the gas
scrubber and connections. Hold for
subsequent analysis.
Williams Cuprous Chloride Procedure
NOTE: It is recommended that this distillation
technique be used for samples containing less
than 1 mg/1 cyanide. In samples of this type
low recoveries have been obtained using the
Serfass Distillation. (See Reference 1.)
II PROCEDURE
A Add 50 ml of 4% NaOH to the gas washer
and dilute with distilled water until the
spiral is covered.
B Add 250 ml of sample to the boiling flask.
Dilute to 500 ml if necessary.
C Connect the train (boiling flask, condenser,
gas washer and trap) to the vacuum line
and adjust the pressure so that approxi-
mately one bubble of air per second enters
the gas scrubber.
D Add 20 ml of mercuric chloride solution
and 10 ml of magnesium chloride solution
through the air inlet tube. Rinse the tube
with distilled water.
E Add slowly 5 ml of cone, sulfuric acid
through the air inlet tube and rinse with
distilled water.
F Heat the sample to boiling, using extreme
care and attention to prevent the sample
from backing up into the air inlet tube.
G Reflux the sample for one hour. Remove
the burner and continue the air flow for
about ten minutes.
I REAGENTS
A 1:1 H2SO4
B Cuprous Chloride
C Sodium Hydroxide, 4% Aqueous Solution
II PROCEDURE
A Add 50 ml of 4% NaOH to the gas washer
and dilute with distilled water until the
spiral is covered.
B Add 250 ml of sample or an aliquot diluted
to 250 ml Claissen flask.
C Connect the train (boiling flask, condenser,
gas washer and trap) to the vacuum line and
adjust the pressure so that approximately
one bubble of air per second enters the gas
scrubber.
D Slowly add 50 ml of 1:1 H2SO4 through the
air inlet tube and rinse with distilled water.
E Add one-half gm Cu2Cl2 to the flask by
washing with distilled water.
*Chemist, DWS&PC Training Activities, SEC.
CH. Cy. lab. Ic. 12.65
24-5
-------
Laboratory Procedure for Cyanide
F Heat the sample to boiling, using extreme
care and attention to prevent the sample
from backing^up into the air inlet tube.
G Reflux for 2 hours.
H After removing heat continue air flow for
about 10 minutes.
I Drain the contents of the gas scrubber
into a 250 ml volumetric flask and bring
to mark with washings from the gas
scrubber and connections. Hold for
subsequent analysis.
Silver-Nitrate and Rhodanine Indicator
(Modified Liebig Method)
D Titrate a 200 ml aliquot of the distilled
sample and record the titration volume.
Save remaining distillate for colorimetric
analysis.
DATA
Blank Standard Sample
ml used _
Corrected
titration value _
mg CN~/aliquot_
mgCN'Aiter
I REAGENTS
A Silver nitrate solution, 0.0192 N.
Dissolve 3. 27 gms of silver nitrate in
1. 0 liter of chloride-free distilled water;
1 ml = 1 mg CN~. Standardize against
standard NaCl using the Mohr method.
B p-dimethylamino benzalrhodanine
solution. Dissolve 0. 02 gms of rhodanine
compound in 100 ml of acetone.
C Stock cyanide solution, 1.0 mg/ml
(1000 ppm).
D Sodium hydroxide, 4% aqueous solution.
II PROCEDURE
A Familiarization with End-Point
Run several blank determinations as
follows: Place 50 ml of 4% sodium
hydroxide and 150 ml of distilled water
in a 500 ml Erlenmeyer flask.
B Add about 10 drops of rhodanine indicator
and titrate with silver nitrate to the first
distinct color change (yellow to pink).
C Repeat the above exercise using 2. 0 or
4. 0 ml of CN" stock solution.
Ill CALCULATION
-/ liter - (ml of AgNO3 - ml for blank) 1000
ml of sample titrated
NOTE: If the above result is less than 1 mg/1,
proceed with the colorimetric determination.
2, 4. A. 1 0 APPARATUS - SCHEMATIC OUTI.INK
K.H. Satgi-nt Calalof; Nos. nml Pru rs Givrn
24-6
-------
Laboratory Procedure for Cyanide
Pyridine-Pyrazalone Method
I REAGENTS
A Chloramine T, 1% aqueous solution.
(Prepare fresh daily. If this does not
dissolve rapidly it indicates Chloramine
T degradation and probable analytical
difficulty.)
B 1-phenyl, 3-methyl, 5-pyrazalone,
saturated aqueous solution. Add 3-4 gms
of the pyrazalone compound to 500 ml of
water and bring to boil. Cool, let stand
overnight and filter the appropriate volume
prior to use. The solution is stable
indefinitely.
C Bis-pyrazalone. Dissolve 0. 025 gm of
bis-pyrazalone in 25 ml in pyridine
(prepare fresh daily). Eastman 6969,
designated as 3, 3'-dimethyl-1, 1'diphenyl-
(4, 4'-bi-2-pyrazalone)-5, 5'dione.
D Mixed Reagent
Immediately prior to use, pour 125 ml of
solution #2 and 25 ml of solution #3
together and mix. (Prepare fresh daily.)
E Acetic acid 1:4
Cyanide stock standard, 1.00 gms
CN~/liter. Dissolve 2. 51 gms potassium
cyanide in 1.0 liter of distilled water.
Add 2 or 3 pellets of sodium hydroxide to
increase stability. Standardize against
silver nitrate and recheck weekly.
II PROCEDURE
A Preparation of Standards
1 Prepare a working standard by diluting
accurately 1. 0 ml of the stock standard
to 1. 0 liter. (This solution is equiva-
lent to 0. 0010 mg CN'/ml).
2 Add 0, 1.0, 2.0, 3.0, 5. 0 and 7. 0 ml
of the working standard to each test
tube and dilute to the 25. 0 ml mark.
B Preparation of Sample
1 Dilute remaining 50 ml of distillate to
about 150 ml. Transfer to a 400 ml
beaker and neutralize with 1:4 acetic
acid using a pH meter. (Caution: Do
not over titrate with acid.) Dilute to
250 ml with distilled H2O.
2 Add 10 ml and 20 ml portions of
neutralized sample to separate test
tubes. Dilute to 25. 0 mark.
DATA
Standard
Sample
Volume Used, ml
0
1
2
3
5
7
10
20
mg CN~ in 25 ml
0
.0010
.0020
.0030
.0050
.0070
--
--
Absorbance Reading
620 mn, 1.0 cm cell
--
--
--
--
--
--
--
--
24-7
-------
Laboratory Procedure for Cyanide
C Color Reaction
1 Add 0. 2 ml of Chloramine T solution
to each standard and sample. Mix and
let stand for 3 minutes.
2 Add 5 ml of mixed reagent to each test
tube, mix and let stand for 20 minutes.
3 Read the absorbance of each sample
and standard in a 1.0 cm cell at 620 mix,
using the O standard as a blank.
D Calculation
1 Construct a standard curve, plotting
mgs CN~ vs optical density.
2 Apply readings from samples to curve
and calculate mg CN~Aiter in the sample.
REFERENCES
Finger, J. H. Recovery of Simple
Cyanides by the Serfass Distillation
Procedure as Compared with the
Williams Cuprous Chloride Method.
Lab. Investigations Report No. 2,
Technical Advisory and Investigations
Section, Technical Services Branch,
Division of Water Supply and Pollution
Control. April 7, 1964.
2 Gonter, C. E. and Schmitt, J. W. Deter-
mination of Cyanides in Water and
Waste Samples. Paper presented before
the Division of Water and Waste Chem-
istry, ACS, New York, New York.
September, 1963.
3 Standard Methods for the Examination of
Water and Wastewater, llth Edition.
APHA, AWWA, WPCF. 1960.
24-8
-------
RECENT ADVANCES IN ESTIMATING FLUORIDES IN WATER
R. C. Kroner*
I INTRODUCTION
Today more than 2, 000 cities and towns in
the United States, serving more than 40
million people, add fluorides to their public
supplies as a caries preventive. For this
purpose, the concentration of fluoride is
maintained at approximately 0. 8 to 1.2 mg/
liter. However, water supplies in many
parts of the country contain naturally occur-
ring fluorides in varying quantities, and in-
dustrial discharges often contribute additional
fluorides to the normal content.
II ANALYTICAL REQUIREMENTS
Since a wide range of skills and experience is
exhibited by the personnel regulating the
fluoridation procedures, a simple, foolproof
method for measuring microquantities in water
is required. The measuring procedure is
frequently preceded by a separation procedure,
usually a distillation, for removal of inter-
ferences. The analytical requirements then
are resolved into:
1 Rapid, dependable separation
procedure
2 Rapid, simple measuring procedure,
III SEPARATION PROCEDURE
A The older separation procedure still in
use'*) is the Willard-Winter steam dis-
tillation. This procedure has the follow-
ing shortcomings.
1 The apparatus used is fragile and
unhandy.
2 Temperature control is extremely
critical because too low a temperature
does not recover the fluoride and too
high a temperature leads to sulfate
carryover from the sulfuric acid dis-
tilling media.
3 Constant attention by the analyst is
required.
B A more recent distillation procedure by
Bellack(2) employs distillation of a sample
from a fixed ratio of sulfuric acid-water
medium, using very simple glassware.
The advantages of this procedure are:
1 Simplicity of equipment
2 Rapid distilling time
3 The procedure does not require
constant attention
4 The sulfate carryover is negligible.
(12)
C A recent method employs the ion-
exchange as a means of separation, using
an ahionic resin. A resin slurry is
pipetted into a column and allowed to
settle and purified sand is added to cover
and preserve the moisture of the resin.
The sample may be poured directly into
the column reservoir - with one exception.
If aluminum is present above 0. 5 ppm,
pretreatment of the sample is necessary.
Pretreatment consists of dosing the sample
with an ammonium-EDTA solution and ad-
justing the pH to 11-12. The fluoride ion
is eluted from the resin by addition of a
beryllium-acetate solution to the column
reservoir. The eluate is collected and
the SPANeS^7) procedure for measuring
fluoride concentration is followed. The
advantages of this method are:
1 Complete elimination of distillation
equipment.
2 The operation is self-sustaining so that
the analyst need not be in close
attendance.
3 Ability to handle multiple samples
simultaneously.
4 Savings in time and equipment.
*In Charge, General Laboratory Services, Water Quality Section, Water Pollution Surveillance
System, 1014 Broadway, Cincinnati, Ohio. Revised by John M. Matthews, Chemist, Analytical
Reference Service, Training Program, SEC. Reviewed December 1965.
CH. HAL. f.33b. 11.64 25-1
-------
Recent Advances in Estimating Fluorides in Water
IV MEASURING TECHNIQUES
A The two Standard Methods in current use
are based upon the complexation of zir-
conium with alizarin dye to form an intense
red solution. When a small amount of
fluoride ion is added, a zirconium-fluoride
complex is formed, thus lessening the
intensity of the red color of the solution.
As the concentration of fluoride increases,
the solution becomes less red and begins
to turn yellow. A point is finally reached
at which the addition of further fluoride
does not appreciably change the color of
the solution.
The older Scott-Sanchis and Lamar
methods were designed for use with Nessler
tubes, i.e., are visual methods. The
Megregian-Maier method used different
concentrations of zirconium and alizarin
and different acidity which adapts the pro-
cedure to photometric measurement of
the color intensity. The disadvantages of
these methods are roughly the same for
each:
1 Sensitivity to interferences, especially
sulfate, phosphates and aluminum
2 Time required for completion of color
reaction, especially with regard to
the Megregian-Maier procedure
B A procedure developed by Megregian
uses a different dye, Eriochrome Cyanine
R, which gives rapid color production.
Procedures are also given to overcome
the interferences due to sulfate and
aluminum. The net result is a photometric
method which gives:
1 Complete color formation in about 15
minutes
2 Built-in steps for removal of two
important interferences
However, since the method is very sensi-
tive to sulfate, this correction step must
be included in most measurements. In
order to circumvent the effect of sulfates.
(6)
Thatcher incorporated barium chloride
into the ECR reagent to precipitate the
unwanted sulfate. This step accomplishes
the desired result, but the time advantage
is lost because of the period required for
growth and settling of the barium sulfate
crystals.
(7)
C A newer method proposed by Bellack
employs a dye known as SPADNS* which
replaces the alizarin dye of the older
methods. The dye solution is incorporated
into the zirconium salt solution so that only
one reagent is required. The advantages
claimed for this procedure are:
180*C
Bellack Fluoride Still
*Eastman Catalogue #7309 (4, 5 Dihydroxy-3-
(p - s ulf ophe nyazo)-2,7- naphthalene - disulf onic
acid, trisodium salt.
25-2
-------
Recent Advances in Estimating Fluorides in Water
1 One shot reagent
2 Color completion in less than 5
minutes
3 Built-in correction for aluminum
interference
4 No pH adjustments required for
sample.
D Other methods which have been proposed
include:
1 A method for turbidimetric titration of
fluoride with thorium nitrate'8)
2 Displacement of silicon from a silicon
fluoride compound and subsequent re-
action of the free silicon to form a
heteropoly blue compound. The amount
of color formed is proportional to the
amount of fluoride in the silicon-
fluoride compound^) and' '.
3 Titration of a fluoride sample with
thorium nitrate and subsequent
measurement of the resulting color by a
recording spectrophotometer.' '
V STANDARD METHODS, llth EDITION
A The Standard Methods which was issued
in 1960 made the following changes:
1 The Willard and Winter steam dis-
tillation was retained as the Standard
Method.
2 The Bellack distillation procedure was
included on a tentative basis.
3 The Lamar visual method was excluded.
4 The Scott-Sanchis visual and the
Megregian-Maier photometric methods
were retained as referee methods.
5 The new SPADNS photometric method
was included on a tentative basis.
TABLE OF INTERFERENCES
Concentration of Substance, In mg/1, Required to Cause
An Error of (+) or (-) 0. 1 mg/1 at 1. 0 mg/1 F
Alkalinity
Al+++
Cl"
Fe+++
(NaP03)g
"PO A "
i\-JA
SO4"
Scott-Sanchis
400(-)
0. 25(-)
2, OOO(-)
2(+)
LOW
300(+)
Megregian-Maier
325(-)
0. 2(-)
1, 800(-)
5(-)
1. K+)
400(+)
SPADNS
5, OOO(-)
0. !(-)*
7, 000(+)
10(-)
1. 0(+)
200(+)
Chlorine Must Be Completely Removed With Arsenite
Color & Turbidity Must Be Removed or Compensated For
*Above Figure is For Immediate Reading. Allowed to Stand Two Hours
Tolerance is 3.0 mg/1. Four Hour Tolerance is 30 mg/1.
25-3
-------
Recent Advances in Estimating Fluorides in Water
REFERENCES
1 Standard Methods for the Examination of
Water, Sewage and Industrial Wastes.
2 Bellack, E. Simplified Fluoride Distil-
lation Method. Jour. AWWA, 50,
April 1958. p 530.
3 Campbell, S. T., Chief Chemist, Torres-
dale Plant. Philadelphia. Unpublished
Data.
4 Shoup, R. Removal of Interferences in
The Scott-Sanchis Fluoride Determi-
nation. Anal. Chem. 29. August 1957.
p 1216.
5 Megregian, S. Rapid Spectrophotometric
Determination of Fluoride with Zir-
conium-Eriochrom Cyanine R Lake.
Anal. Chem. 26. 1954. pllGl.
6 Thatcher, L. L. Modified Zirconium
Eriochrome Cyanine R Determination
of Fluoride. Anal. Chem. 29. November
1957. p 1790.
7 Bellack, E., and Schouboe, P. Rapid
Photometric Determination of Fluoride
with SPANDS. Zirconium Lake.
Unpublished.
8 Brandt, W. W., and Duswalt, A. A. Determi-
nation of Fluoride Ion by Turbidimetric
Titration. Anal. Chem. 30. June 1958.
p 1120.
9 Curry, R.P., and Mellon, M. G. Spec- '
trophotometric Determination of In-
organic Fluoride and of Fluorine in
Organic Compounds. Anal. Chem. 29,
November 1957. p 1633.
10 Curry, R. P., and MeUon, M. G. Colori-
metric Determination of Fluoride in
Water by Heteropoly Blue System.
Anal. Chem. 28. October 1956. p 1567.
11 Mennis, O., Manning, D. L., and Ball,
R. G. Automatic Spectrophotometric
Determination of Fluoride. Anal.
Chem. 30. 1958. p 1772.
12 Kelso, F. S., Matthews, J. M., and
Krame r, H. P. Ion-Exchange Method
for Determination of Fluoride in Pot-
able Waters. Anal. Chem. 36:577.
March 1964.
ADDITIONAL REFERENCES
1 Kramer, H. P., Kroner, R., and Ballinger,
D. G. Problems in Estimating Fluorides
in Water. Jour. AWWA 48. May 1956.
p 573.
2 Thorn, J. V., and Gribkoff, G. P. Com-
parative Fluoride Analysis by Several
Methods. Jour. AWWA 48. April 1958.
p 455.
25-4
-------
LABORATORY PROCEDURE FOR FLUORIDE
John M. Matthews*
REAGENTS
A Resin: Purification
To purify the resin,** wash in the following
manner and decant after each wash. Place
450 grams of resin in a 1500-ml beaker,
wash with two 300-ml portions of 95%
ethanol; two 300-ml portions of distilled
water; five 300-ml portions of 3 M hydro-
chloric acid; then transfer to Sedwick
Rafter cone and wash with 8 liters of dis-
tilled water by means of the siphon arrange-
ment. Transfer resin to 1500-ml beaker,
add 300 ml of 1 M sodium acetate solution
and stir with magnetic stirrer for 15
minutes. Discard supernatant. Prepare
a 1:1 slurry of the acetate-resin and dis-
tilled water and store in a closed poly-
ethylene container.
B Beryllium Eluting Solution - Stock
Solution
Pipet 57 ml of glacial acetic acid into 500
ml of distilled water, dissolve 2.6 grams
of reagent grade beryllium carbonate
(BeO)5-CO2'5H2O*** and dilute to 1 liter
with distilled water. (Note toxicity of
beryllium, and exercise care.)
C Beryllium Eluting solution - Working
Solution
The beryllium stock solution is diluted
1:10 with distilled water to provide the
beryllium eluting solution used to remove
the fluoride from the acetate resin.
D Sand: Purification
Use purified white Quartz sand,**** 60-
120 mesh. Purify the sand by digestion
at 100°C for 1 hour with 250 ml of 20%
sodium hydroxide, discard supernatant,
and wash with 250 ml of 1:3 hydrochloric
acid solution. Wash in 2-liter flask with
distilled water until all traces of chloride
are removed.
E Ammonium - EDTA Solution
To prepare the 0, 13 M NH4EDTA solution,
dissolve 50 grams of Na2C10H14O8N2'2H2O
in 70 ml of concentrated NH4OH and dilute
to one liter with distilled water.
F SPADNS Reagent
Dissolve 0.958 grams 4, 5-dihydroxy-3-
(p-sulfonphenylazo) -2, 7-naphthalene
disulfonic acid trisodium salt (Eastman ,
No. 7309) in distilled water and dilute to /
500 ml.
G Acid-Zirconium Reagent
Dissolve 0. 133 grams zirconyl chloride
(ZrOCl2 • 8H2O) in 25 ml distilled water.
To the zirconium solution add 350 ml
cone. HC1 and dilute to 500 ml.
Combine equal volumes of the above re-
agents to produce a single working reagent.
H Reference Solution
Mix 10 ml of SPADNS reagent solution
with 103 ml of distilled water and 7. 0 ml
of cone. HC1.
I Fluoride Stock Solution
Dissolve 0.2210 grams NaF in 1 liter of
distilled water (1 ml = 0. 1 mg F).
J Standard Fluoride Solution
Dilute 100 ml of the above stock solution
to 1 liter (1 ml = 0.01 mg F). From this
prepare working solutions.
**Dowex 2-x8, Anionic Resin, Cl Form,
50-100 Mesh. Dow Chemical Co.
***Fisher Scientific Company
#***Fisk Sand Company, 40-38th Ave. North,
Minneapolis, Minnesota
*Chemist, Analytical Reference Service, Training Program, SEC. Reviewed December 1965.
CH. HAL. f. lab. 1. 11.64 25-5
-------
\
Laboratory Procedure for flucO&de •-
^i
II PROCEDURE
( A Ion Exchange
/.
1 Preparation of columns for each
standard and sample.
a While the magnetic mixer is stirring
and purified resin slurry, pipet 25ml
into each reservoir. After the resin has
settled in the columns, rinse the remain-
ing resin from sides of the reservoir.
b Add purified sand to each column to
form a top layer approximately 1 cm
high. Wash the reservoirs with
distilled water to remove all sand
from the sides of the reservoirs.
Allow the water to drain completely -
discard the drainage.
2 Analysis of water sample
a If no aluminum is present in the
sample then proceed to step b. If
alumium is present in excess of
0. 5 mg/1 then proceed as outlined
below.
1) Pipet 50 ml of the standard
fluoride working solution, (. 00
mg, .02mg, .04mg, .06mg/50
ml) into individual 125-ml Erlen-
meyer flasks.
2) Pipet 50 ml, or an aliquot diluted
to 50 ml, of sample into a 125-ml
Erlenmeyer flask.
3) To each 50 ml volume of standard
and sample pipet 1 ml of NH4EDTA
solution followed by 2 ml of 0. 5
N4NaOH. Mix well.
4) Pour each standard and sample
into its respective prepared
column and allow to drain. Dis-
card the drainage. Proceed to
step d.
b Pipet 50 ml of the standard fluoride
working solution into its prepared
column.
c Pipet 50 ml, or an aliquot diluted
to 50 ml, of sample into the pre-
pared columns and allow to drain.
Discard the drainage.
d Wash the columns by adding 100 ml
of distilled water to the reservoirs
and allow to drain. Discard the
drainage.
e Remove the adsorbed fluoride from
the resin by pipetting 100. 0 ml of
beryllium eluting solution into the
reservoirs. Collect the eluate in
a 250-ml Erlenmeyer flask.
B Colorimetric (SPADNS)
1 Pipet 20 ml of mixed SPADNS solution
into each flask containing standards
and samples. Mix well.
2 Adjust the temperature of the standards
and samples to within + 2°C of each
other.
3 Read the standards and samples in 1 cm
cells against the reference solution at
/ iO -5*fr mji. If aluminum was present in
(0 the original sample and step 2a was
used for complete complexation of the
aluminum fluoride complexes, then a
30-minute waiting period is required
for a complete color development.
4 Prepare a standard curve, plotting
mg F vs. OD.
25-6
-------
TRACE ORGANIC CONTAMINANTS IN WATER
R. L. Booth*
I INTRODUCTION
The subject of trace organic contaminants
in water continues to receive ever-increasing
amounts of attention. Concurrently, the
sources of these refractory materials are
becoming more varied and complex. The
problems associated with these substances
are, likewise, increasing and satisfactory
water treatment is becoming more and more
difficult.
II SOURCES
A Man-made
i^l Domestic wastes, in various stages of
sewage treatment, discharged into
rivers and streams.
^ 2 Industrial wastes, due to their multi-
plicity and complexity.
3 Carrier solvents, such as those used
in pesticide formulations.
4 Chemicals applied directly to land
and water.
B Natural
•/'l Extracellular products of algae by
(a) diffusion of metabolic intermediates,
(b) by-products of metabolism, and
(c) hydrolysis of capsular materials.
i/ 2 Actinomycetes, microorganisms present
in rivers/streams, by their growth and
decomposition cycles. ( j<2.
-------
Trace Organic Contaminants in Water
where large quantities of organic
materials are needed.
B Isolation
1 The carbon samples are sequentially
extracted with chloroform and ethyl
alcohol to desorb the organic material
from the carbon.
a After extraction, the excess solvent
is removed, and the samples are
brought to dryness to yield a:
1) Carbon chloroform extract (CCE).
2) Carbon alcohol extract (CAE). O*
2 The CCE is separated into broad
classical groups by techniques based
on solubility differences.
3 Further separations are made by such
techniques as adsorption, paper, and
thin layer chromatography.
C Identification
1 Tentative identification is normally
made by gas chromatography analysis.
2 Positive identification normally
requires infrared confirmation.
VI TREATMENT AND REMOVAL PRACTICES
A Plant treatment procedures, such as
coagulation, sedimentation, and filtration
are generally not too effective.
B Chemical treatment
1 Copper sulfate used to control algae.
2 Oxidizing agents, such as chlorine,
chlorine dioxide, ozone, and potassium
permanganate are used with varying
degrees of success.
3 Activated carbon treatment removes
organic substances by adsorption.
C Biological Treatment
1 Natural degradation in unsaturated
soils and streams.
2 Biological oxidation of organic
materials both in streams and
acclimated systems.
VII SUMMARY
The problems associated with trace organic
contaminants in water are becoming more
V •* apparent as our need and usage of water
increase. The continued growth of the
chemical industry, our increasing population,
and the public's demand for more palatable
water emphasize even more the urgency of
these problems. Developments have been
made in the detection of these refractory
materials and in their removal from water
supply sources. It is apparent, however, that
further advances in the collection, identification,
and removal of these pollutants are needed to
insure the public of high-quality water and
water resources.
REFERENCES
1 Anon. Tentative Method for Carbon
Chloroform Extract (CCE) in Water.
Jour. AWWA. 54, 223. 1962.
2 Ryckman, D. W., Burbank, N. C.,Jr., and
Edgerley, E., Jr. New Techniques for
the Evaluation of Organic Pollutants.
Ibid. 56, 975. 1964.
3 Ettinger, M. B. Developments in Detection
of Trace Organic Contaminants. Ibid. 57,
453. 1965.
4 Robeck, G. G., Dostal, K. A., Cohen, J. M. ,
and Kreissl, J. F. Effectiveness of
Water Treatment Processes in Pesticide
Removal. Ibid. 57. 181. 1965.
5 Anon. Taste and Odor Control - Chemicals
and Methods. Taste and Odor Cont.
Jour. 31. No. 1, 1. 1965.
26-2
-------
METHODS OF RECOVERING ORGANIC MATERIALS FROM SURFACE WATERS
R. H. Burttschell and J. J. Lichtenberg*
Methods for recovering organic materials
from water may be classified according to
the degree of concentration required before
the desired analytical procedure can be
applied.
I CONCENTRATED SOLUTIONS
Where only a minor degree of concentration
is involved, the following methods should be
considered.
A Liquid-liquid extraction usually involves
water and an immiscible organic solvent.
Solvents should be investigated in the
series of increasing polarity:
aliphatic hydrocarbons
aromatic hydrocarbons
ethers
chlorinated compounds
esters
alcohols, amines, acids, etc.
pH may be of critical importance; this
point will be discussed at length in the
discussion of Analytical Procedures. In-
organic salt concentration may also be
important.
Remember the concept of the partition
coefficient:
K =
Cs
Cw
where K is the partition coefficient
Cs is concentration in the extract-
ing solvent
Cw is the concentration in water
Continuous extractors must be used where
the K value is not favorable to the extract-
ing solvent. Continuous batch and counter-
current extractors may be used.
Separatory funnels are available in a variety
of sizes. Batch continuous extractors are
commercially available in sizes up to a
liter or so and can be readily assembed in
larger sizes. Continuous counter-current
extractors are convenient for up to 10-20
gallons.
B Steam distillation can be used to strip a
sample of small amounts of volatile or-
ganic compounds; in this case the distillate
usually must be further concentrated by
liquid-liquid extraction. The organic com-
pound should have at least moderate vapor
pressure at 100 C and should be almost
insoluble in water.
A variation of this method is simple
evaporation to concentrate non-volatile
organic compounds.
C Precipitants may be used: silver salts of
acids, chloroplatinates or tetraphenyl
boron, derivatives of amines, phenylhy-
drazine derivatives of ketones and alde-
hydes, etc.
D Ion exchange may be used to concentrate
acids and bases. The only limit to the
volume of water to be filtered is the
amount of inorganic salts in the water;
these inorganic salts usually use up the
exchange capacity too rapidly to make this
method practicable.
II VERY DILUTE SOLUTIONS
To concentrate extremely dilute solutions the
carbon filter is the most useful method. The
organic matter is adsorbed from aqueous so-
lution and desorbed by an organic solvent.
The great advantage is the large amount of
water that can be put through a small filter;
the disadvantages lie in the lack of quantita-
tive adsorption and desorption.
A The Adsorption Process
1 The adsorption process involves an
equilibrium in solution.
Adsorbed
.Unadsorbed
*Chemist, Chemistry & Physics Section Section, Basic and Applied Sciences Branch, DWS&PC,
SEC and Chemist, Water Quality Section, DWS&PC, SEC. Reviewed December 1965.
CH. OTS. 37a. 3.62
27-1
-------
Methods of Recovering Organic Materials from Suv .ice Waters
2 The Freundlich Isotherm is often useful
in evaluating adsorption behavior.
1
x
m
= kc
Units may be chosen for convenience.
The larger _ the stronger the adsorp-
tion. The n constant k gives the
— value at unit concentration.
m
x = weight of adsorbed compound
m = weight of carbon
k = constant
c = equilibrium concentration of
organic compound in the liquid
phase
— = constant
3 General rule: non-polar compounds are
strongly adsorbed and polar compounds
weakly adsorbed from water e. g. , mi-
meral oil is more strongly adsorbed than
glycine.
4 Adsorption will depend on the type of
carbon, grain size, pH of solution, po-
larity of adsorbate, nature of the sol-
vent, temperature, etc. If powdered
carbon is not used, the column length
and contact time must be chosen so as
to permit efficient adsorption.
5 Desorption is simply the reverse re-
action in the adsorption equilibrium
and is influenced by the same factors
as adsorption. It is usually necessary
to use an organic solvent for desorption
with an extended period of extraction.
B Setting up the Carbon Filter
1 In the next section is a description of
the procedure used in setting up and
running carbon filters in our laboratory.
It should, however, be understood
that elaborate apparatus is not required
and that excellent results can be ob-
tained with a piece of glass tubing
closed at either end by a rubber stopper
in which is inserted a piece of smaller
glass tubing. Such apparatus is ade-
quate for testing small volumes, e. g. ,
sewage or concentrated industrial wastes
although not convenient for handling
thousands of gallons of water.
Wherever the investigation is concerned
only with certain specific compounds,
bench tests in small columns should be
carried out in order to evaluate the per-
formance to be expected from the filter.
TTI INSTALLATION OF EQUIPMENT AND
COLLECTION OF CARBON FILTER
SAMPLES
A The carbon filter consists of a piece of
pyrex glass pipe 3 inches in diameter and
18 inches in length. The ends are fitted
with brass plates and 3/4 inch galvanized
nipples. A stainless steel screen is
fixed in a neoprene gasket at both ends.
B Presetting, Prefilter, and Backwash
River waters will frequently clog the car-
bon filter before the desired amount of
water has been sampled. It is necessary
to remove sufficient turbidity to permit
the required amount of water to pass
through the unit. This may require a pre-
settling tank, and a prefilter containing
sand and gravel. For waters having less
than 100 ppm of turbidity, a presettling
tank is generally not required. Tap water,
of course, requires no prefiltering and
may be passed directly into the carbon
filter.
C Presettling Tank
A standard hot water tank connected with
the inlet at the bottom and outlet at the top,
with a clean-out tap at the bottom can serve
as a presettling tank. The outlet connects
to the prefilter containing sand and gravel.
The hot water tank should be flushed at fre-
quent intervals to prevent a large accumu-
lation of solids. Open settling tanks can
be used if care is taken to prevent long
detention times and biological action. With
open tanks a pump will be required to move
the water through the filter.
-------
Methods of Recovering Organic Materials from Surface Waters
D Sand Prefilter
The sand prefilter consists of a standard
piece of steel pipe 3 inches in diameter
and 3 feet long, threaded at both ends, and
equipped with 3" X 1" reducer couplings.
Two disks of stainless steel screen are
fitted to the inside diameter of the pipe.
A simple way to prepare and hold the screen
in place is to cut out a disk about the size
of the outside diameter of the reducer,
and then push it into place so that it is be-
low the threads. The screen is held
tightly against the inside of the reducer.
The pipe is packed with 6 inches of 1/8"
gravel, 24" of 0. 6 to 0. 8 mm sand, and
another 6 inches of 1/8" - 1/4" gravel.
No free space is left in the pipe. The
gravel should be packed by jarring the
pipe while fitting. This arrangement
provides a strainer rather than a filter
with a movable bed. By such an arrange-
ment backflushing can be done without
disturbing the filter. The construction
details are shown in Figure 1. The di-
mensions may be varied according to
local conditions.
IV FILTER ARRANGEMENT
A The presettling tank, the sand prefilter,
and the carbon filter should be installed
at the most convenient source of raw water.
If less than 15 psi pressure is available,
it may be necessary to pump the water
through the system. A schematic drawing of
a workable system is shown on the following
page. Exact lengths of pipes, etc. are
not given, since these will vary with the
16-18 Mesh
Stainless Steel
Screen •
J~U-
Standard 3" Pipe
Threaded Each End
16-18 Mesh
Stainless Steel
Screen
" X 1" Reducer Coupling
lo l/8"-l/4" Gravel
0.6 to 0. 8 mm Sand
1/8" - 1/4" Gravel
3" X 1" Reducer Coupling
FIGURE 1 - Details of Sand Prefilter
27-3
-------
Methods of Recovering Organic Materials from Surface Waters
local situation. Both the sand and car-
bon filters should be connected with unions
at both ends for easy removal. A typical
installation is shown in the photograph on
page 3-6.
B The raw water passes into the bottom of
the sand prefilter, around and into the
bottom of the carbon filter. When the rate
of flow through the system falls below
1/4 gpm backwashing of the sand filter is
necessary, using a high pressure source
of water. A clean hose is connected to
the top of the sand filter, the valve to the
carbon filter is closed, the drain valve
on the sand filter is opened and the sand
is back-flushed until the water coming out
is clear. The length of time between '
backwashings will vary. On the Missouri
River, for example, it has been found
that once every 24 hours is sufficient.
After backwashing connect the system as
before and continue the sampling. Be sure
to disconnect hose at top of sand filter
after backwashing. A pressure gauge is
inserted in the system to indicate when
clogging is taking place in the carbon fil-
ter. Total pressure in the filter should
not exceed 50 psi.
C A water meter, located at the end of the
system to prevent excessive fouling of
the meter, is used to measure the vol-
ume of water samples. It is good practice
r
Union-
Hose Connection
j Valve
Valve
Sand Prefilter'
Pump
(If Required)
Raw
Water
Water
Meter
•Carbon Filter
Pressure ^ j2^Uni
Gauge
LJl
Valve.
Drain-
•Flow
Regulator
(1/2 GPM)
Figure 2 - Schematic Diagram of Piping Installation for Sand
Prefilter and Carbon Filter
27-4
-------
Methods of Recovering Organic Materials from Surface Waters
• to disassemble and clean the meter thor-
oughly after each run. A 5/8" X 3/4",
disk-type meter, is fairly satisfactory.
A valve following the meter serves to
throttle the flow if necessary. A flow-
regulating device may also be used for
this purpose. The complete installation
is shown in the schematic diagram,
Figure 2.
D Fine carbon washes out of the filter when
it is first started. A few gallons of water
can be passed through the top connection
and through the carbon filter drain before
cutting in the meter, to keep the meter
free of the carbon. The hose connection
and drain on the carbon filter can be used
to back-flush the carbon should it become
clogged. This is a last resort and should
only be used if absolutely necessary.
E The system outlined is not intended to re-
move all traces of turbidity from the water
before passing through the carbon filter.
Its purpose is to take out gross materials,
most organisms, and permit the required
volume of water to pass through the car-
bon. The valve nearest the bottom of the
sand filter is to be closed when it is desired
to obtain a sample of raw water directly
for other analyses. The drain valve is
opened and, after flushing, the necessary
raw water sample is collected.
F Pumping
1 If adequate pressure is not available
for sampling raw water, it is neces-
sary to pump through the filter system.
It is important that the pump used
should not contaminate the sample
through oily packing or other sources.
New pumps are sometimes grease
coated and should be thoroughly cleaned
before being put into service. If a lift
of more than 12 feet is required, it
will be necessary to use a jet-type
pump. A model suitable for the con-
ditions should be selected.
2 Before a new pump is put into service,
it should be thoroughly flushed with
hot water containing a little detergent.
The pump should be operated against
the minimum, amount of head required
to get the water through the filter. This
may require by-passing of part of the
flow.
G Collection of Sample
1 With fairly concentrated samples (sew-
age, industrial wastes, etc.) a small
filter and a few gallons of water may
be sufficient. Generally, for river
sampling, water should be passed
through a large filter at a rate of 1/4
to 1 / 2 gallons per minute until up to
5000 gallons or more have been filtered.
With highly turbid waters clogging may
occur earlier. Although a suitable sam-
ple can sometimes be obtained with
several hundred gallons of river water,
it is desirable to filter a minimum of
2000 gallons if at all possible. A flow
regulating device set at 1/4 or 1/2
gpm can be placed in the system ahead
of the carbon filter. A suitable unit can
be obtained from the Dole Valve Company,
6201 Oakton Street, Morton Grove,
Illinois. These devices generally need
a minimum pressure of 15 psi for proper
operation.
2 Since the purpose is to get a quantitative
measure of organics in water, it is very
important to have accurate flow measure-
ments. If difficulties in flow occur this
should be noted on a log sheet. Meter
readings should be recorded daily on
log sheets and should be designated
either in GALLONS or CUBIC FEET.
If a satisfactory meter cannot be ob-
tained, the flow rate for a set volume
(1 or 2 liters) can be determined at
regular intervals.
H Precautions
The purpose of the carbon filter is to ad-
sorb small amounts of impurities from
the water. It is important to avoid contami-
nation of the carbon from other sources.
Hence the following should be observed:
1 New galvanized fittings are usually
coated with oil or grease. The oil
should be removed by washing in kero-
27-5
-------
Methods of Recovering Organic Materials from Surface Waters
V
sene followed by a detergent wash before
such fittings are used for making con-
nection to the filter.
2 Ordinary organic pipe jointing com-
pounds should not be used. Red lead
(lead oxide) mixed to a paste with water
can be used for this purpose.
3 Plastic hose is to be avoided, and if
rubber hose is used in any connections
it should be flushed thoroughly before
being connected to the filter. Copper
tubing is ideal for connections.
After the required volume of water has
been run through the filter, the carbon is
removed, dried, and extracted.
The wet carbon is spread out in a thin layer
on a metal sheet and air dried for several
days. The time necessary for drying in-
creases with the thickness of the layer.
Warm air can be passed over the surface
of the carbon to hasten the drying but with
the risk of driving off weakly held volatile
substances.
LABORATORY TREATMENT OF SAMPLES
OBTAINED BY THE CARBON ADSORPTION
TECHNIQUE: EXTRACTION PROCEDURES
The sample is collected by passing approxi-
mately 5000 gallons of water through a carbon
adsorption unit. The unit is shipped to S. E. C.
along with a daily record of the sampling activi-
ty and the carbon treated as described below.
A Preliminary Treatment
1 Records
a Log sample in - list date received -
assign it a number.
b Send daily record sheet to the Lab-
oratory where pertinent data are re-
corded on a data card, i. e. , source,
location, dates sampled, date re-
ceived and total flow.
2 Drying
a Remove wet carbon from the tube and
dry it on copper, brass or stainless
steel trays in an oven at 40°C. (N. B.
air circulated through drying cabi-
net should be prefiltered through *
carbon to prevent the adsorption of
foreign materials). Drying requires
about two days.
b Store carbon samples in one gallon
paint cans, (tightly closed)
B Solvent Extraction of Carbon
It is necessary that blanks be run on all
solvents and on the carbon used for col-
lection of the sample.
1 Packing the Extractors
a Filter paper at bottom of soxhlet.
b Glass wool (pre-extracted) three
inches in depth to prevent carbon
fines from passing into the pot. Add
chloroform to wet the wool.
c Carbon is added and packed. (N. B.
do not pack too tightly)
2 Chloroform Extraction
a Add chloroform - about two cylinder
volumes.
b Extract for 35 hours.
c Siphon and blow the bulk of the chloro -
form into the pot with air.
d Remove the flask containing the
chloroform solution from the system.
Concentrate to 250 ml. by distillation
and filter into a 300 ml. Erlenmeyer
flask. Evaporate to about 20 ml. on
a steam bath with a stream of air and
transfer to a tared 5 dram vial. The
remaining solvent is evaporated at
room temperature without an air jet
and the weight is obtained.
3 Alcohol Extraction
a Remove residual chloroform from
the carbon by one of the following
methods:
27-6
-------
Methods of Recovering Organic Materials from Surface Waters
1) Blow warmed (40°C) prefiltered
air upward through the carbon for
about 4 hours.
2) Leach the residual chloroform off
the carbon with ethyl alcohol (95%)
and distill off the chloroform
alcohol mixture. Repeat until
virtually pure alcohol remains.
3) Remove the carbon from the
soxhlet and air dry on trays.
When dry, repack into soxhlet as
before (Chloroform vapors must
be disposed of).
b Add sufficient ethanol (about two
cylinder volumes) and extract for
24 hours.
c Concentrate the alcohol solution as
in the case of chloroform, except
that the final drying can be carried
out on a steam bath with a stream
of air.
The method listed first is most satis-
factory for our work here, mainly
because a minimum of supervision is
required and chloroform vapors are
exhausted out the hoods.
C Extractable Materials
1 Calculation of concentrations
On most waters it is convenient to
compute the recovery in parts per
billion (ppb)(i. e., (ig/1) using the
following formula.
/?
h - grams recovered X 10
PP " gallons filtered X 3. 785
2 Infrared spectra
Infrared spectra are routinely run on
both the chloroform and alcohol extracts.
D Special Applications
Other solvents may find use in performing
extractions. If special applications are
needed, a testing program is necessary
to establish what solvents may best be
used.
E Adsorption and Desorption
The effectiveness of adsorption and de-
sorption varies for different materials and
should be considered in interpretation of
results.
27-7
-------
PRELIMINARY SEPARATION OF EXTRACTS
R. H. Burttschell*
I INTRODUCTION
As a general principle it may be stated
that the more sensitive modern analytical
techniques (gas and other forms of chroma-
tography, spectrophotometry, etc. ) require
fairly pure samples to begin with; and also
as a general principle, it may be said that
the more widely different the purification
steps employed, the greater the degree of
purification.
Ion exchange, dialysis, crystallization,
electrophoresis, etc. , are useful but cannot
be discussed here; we will briefly cover
only three techniques.
A Distillation
B Partition
C Adsorption
II THEORETICAL BACKGROUND
layers are present, irrespective of
their relative amounts, the total vapor
pressure remains constant, and the
system boils at a definite temperature."
Besides the familiar case of steam
distillation, non-polar organics can
be co-distilled with polar liquids, e. g. ,
insecticides with glycerol, and polar
compounds with non-polar solvents,
e. g. , mineral oil. The co-distillation
can be done with superheated solvents
and under vacuum. Where trace
quantities of one component are in-
volved, there may not be a two-phase
system present and the solvent vapor
serves only as a carrier gas. However,
here the solute can usually be easily
recovered so the method is worth con-
sideration, regardless of the mechanism.
In practical work the three important
points are the stability of the sample,
its volatility, and the ease of recovery
from the distillate.
A Distillation
1 Raoult's Law states that the partial
pressure of A (PA) m an ideal mixture
of volatile solvents is
PA = mole fraction of A in liquid X
vapor pressure of pure A at the
temperature of the system.
The total vapor pressure (which of
course sets the boiling point) is then
PA+PB+ . . . This is the basis
for studies of distillation. The ques-
tion of non-ideal solutions is too com-
plicated to discuss. Most physical
chemistry texts go into it at length.
2 Partially miscible liquids are a
special case, often discussed as
"steam distillation" although "co-
distillation" is a better term. Quoting
from Glasstone, "As long as the two
3 Distillation from acid and basic solu-
tions may also be useful. The principle
behind this will be discussed in the
solubility separation method.
B Partition
Partition is the principle utilized in
liquid-liquid extraction, paper chroma-
tography and gas chromatography. The
fundamental law is the Distribution Law:
"A dissolved substance distributes itself
between the layers of a two-layer system
so that at constant temperature the ratio
of the concentrations is also constant.
The Law is properly applied only to dilute
solutions but a first approximation to the
distribution ratio can be obtained from
the ratio of the solubilities:
K -
Sw
-Chemist, Chemistry & Physics Section, Basic and Applied Sciences Branch, DWS&PC, SEC.
Reviewed December 1965.
CH.OTS.46a.4.65
21-1
-------
Preliminary Separation of Extracts
K is a true constant but apparent dis-
crepancies occur where the solute has
different molecular weights in the two
phases. Benzole acid, for instance, is
largely tjiCOOH plus some benzoate ion
in water; in benzene it is the dimer
(4>COOH)2. K then applies only to the
species common to both layers, COOH.
and ignores the dimer.
Most texts on qualitative organic analysis
contain a great deal of practical infor-
mation on solubilities, while the usual
undergraduate physical chemistry texts
cover the essentials of the theory.
C Adsorption
There are two kinds of adsorption to be
considered, physical (van der Waals)
adsorption and chemisorption. The
theory of adsorption from solution is
not as well understood as adsorption of
gases but we can make a few comments.
Physical adsorption depends on the
relatively weak van der Waals forces due
to electrostatic attraction of dipoles, it
being assumed that non-polar molecules
act as oscillating dipoles, i. e. , the
nuclei of atoms and molecules form
oscillating dipoles with the electrons.
Since the bonds are weak, adsorption can
be made reversible under proper conditions.
Chemisorption is thought of as actual
chemical reaction and the resulting bonds
are much more difficult to break. Losses
due to "irreversible adsorption" may
therefore appear; this is particularly
true of carbon, less so of silica and
alumina.
Ill SOLUBILITY SEPARATION OF
EXTRACTS
In the next section we have described a
laboratory procedure we have found very
useful in handling carbon filter extracts
( CCE). The extracts are split into acid,
basic, and neutral fractions and the neutral
fraction is further separated by adsorption
chromatography.
This procedure has been extremely useful
where the fractions are weighed and infrared
spectra made. However, where volatile com-
pounds are to be looked for by gas chromato-
graphy, it is almost imperative that a steam
distillation step be included. Otherwise the
mass of heavy probably polymeric material
present causes much interference.
Really good clean-up methods are likely to
be highly specific and need to be "tailor-
made" to suit the situation. Since this lecture
is concerned only with "preliminary" puri-
fication, we have omitted discussion of precise
analytical partition and adsorption columns;
thin-layer, ion exchange, paper, and gas
chromatography; gradient elution methods,
etc. Such methods constitute a second and
higher degree of purification but ordinarily
require that they themselves be preceded by
a "rougher" primary purification.
IV NOTE ON LOSSES OF VOLATILE
COMPOUNDS
It may not be generally recognized but it is
quite true that heavy losses of even moderate-
ly volatile compounds occur if one tries to
evaporate off all the solvent, as when pre-
paring to weigh an extract. We have had
losses of over 50% in attempting to carry
out what we thought were very careful evapora-
tions of ether from 10 to 20 mg amounts of
phenols.
For this reason we suggest that no extract or
fraction be evaporated down for weighing
unless it is known to be completely non-
volatile. This precaution becomes more
necessary as the purity of the sample in-
creases because non-volatile impurities act
as "solvents" to reduce the amount of loss
(according to Raoult's Law).
27-9
-------
PROCEDURES FOR THE PRELIMINARY SEPARATION OF EXTRACTS
R. H. Burttschell and J. J. Lichtenberg*
I STEAM DISTILLATION OF EXTRACTS
Steam distillation need be only a very simple
procedure. Put a gram or two of crude or
partially purified sample in a small flask
containing 50 - 100 ml of distilled water.
Set up for ordinary distillation and boil off
most of the water. If necessary, add more
water to the flask and continue distilling un-
til the distillate has no trace of cloudiness or
a second phase as it drips from the condenser.
Transfer the distillate to a separatory funnel,
extract with a suitable solvent, and concen-
trate to a convenient volume. As the product
is relatively volatile, do not try to evaporate
to dryness if it can be avoided. The non-
volatile residue in the distillation flask can
also be recovered if necessary.
If it is desired to recover acids (e. g., phenols
in the 4-AAP colorimetric method), make the
solution acid before distilling; the distillate
will then contain only neutral and acidic sub-
stances. The neutrals can then be separated
during the extraction step. Basic materials
remaining in the still pot, e. g., aniline, can
then be recovered by making basic and dis-
tilling over a second fraction.
II SOLUBILITY SEPARATION OF EXTRACTS
A The organic contaminants from water as
extracted from carbon or concentrated by
liquid extraction ordinarily form an ex-
ceedingly complex mixture for which there
is no one satisfactory separation procedure.
If the analysis is directed towards a single
component, the procedure may be designed
for that purpose; otherwise a useful and
generally applicable preliminary separa-
tion may be made on the basis of relative
acidities.
B
By extracting an ether solution of the sam-
ple with water, then with dilute hydrochlo-
ric acid, then sodium bicarbonate, and fin-
ally sodium hydroxide, a separation into
water soluble, basic, strongly acidic, weak -
ly acidic, and neutral fractions may be made.
The portion insoluble in ether may also be
recovered as an additional fraction. This
method is obviously not suited to very vola-
tile substances, nor to further separation
of the water soluble fraction, nor to sub-
stances unstable in the presence of water
or dilute acid or base; in addition, it must
be remembered that substances whose par-
tition coefficients are not extremely un-
favorable to water may be present in sev-
eral fractions in varying amounts.
C This procedure constitutes one of the best
preliminary steps in analyzing any unknown
sample and may often profitably be either
preceded or followed by steam distillation
(at various pH's) fractional crystallization,
etc. Simple micro qualitative tests for ni-
trogen, halogens, sulfur, phosphorus, etc,
are also often useful; for a more extended
discussion, see the texts mentioned in the
bibliography.
D General
The sample should be substantially free of
solvent. The amount may vary consider-
ably but one-half gram is a very convenient
amount; as little as 100 mg or even less
may be used, but in this case there may
be large percentage errors. Ethyl ether
has proved to be an excellent solvent al-
though often not all the sample will dis-
solve in it; benzene or chloroform may be
used although these solvents are more
likely to cause troublesome emulsions.
Ill LABORATORY DIRECTIONS
A Solubility Separation Procedure
Solution in Ether
1 Weigh previously dried sample con-
tained in a tared 50 ml beaker (ap-
proximately 0. 5 gm preferred). Also,
weigh a 125 ml flask for the Water
Solubles (WS).
*Chemist, Chemistry & Physics Section, Basic and Applied Sciences Branch, DWS&PC, SEC
and Chemist, Water Quality Section, DWS&PC, SEC. Reviewed December 1965.
CH. OTS.22b.4.65 2*7-10
-------
Procedures for the Preliminary Separation of Extracts,
2 Dissolve the sample with a small
amount of methanol (about one ml).
Add the methanol dropwise and stir
with a rigid wire until the sample
reaches a syrupy consistency.
3 Add 30 ml ether to the sample. Not
all will dissolve. Collect the Ether
Insolubles (El) on a sintered glass
funnel with suction and set aside for
paragraph 4. Pour the contents of
the suction flask into a 125 ml separa-
tory funnel equipped with a teflon stop-
cock. (Stopcock grease will contami-
nate the sample.)
Ether Insolubles (El)
4 Dissolve "El" previously collected on
the sintered glass funnel with methanol.
Wash funnel once with a small volume
of chloroform and collect in the suction
flask. Transfer filtrate to the original
tared beaker with methanol. Evaporate
to dryness on a steam bath and record
weight for Ether Insolubles.
Separation of the Water Solubles (WS)
(3>
5 Shake the ether solution three times
with 15 ml portions of distilled water.
Drain the water layers into the weighed
"WS" flask after each shaking. Evapo-
rate the Water Soluble fraction to dry-
ness on a steam bath with -a jet of clean,
dry air and record weight for Water
Solubles.
Separation of Amine Fraction (B)
6 Mark flask "HCl". Shake ether solu-
tion three times with 15 ml portions of
5% HCl and drain aqueous layers into
HCl flask after each shaking. Make
this aqueous solution strongly basic
by carefully adding NaOH pellets
(about 30) or 25% NaOH. The solution
becomes darker at this point. Set a-
side for back-extraction.
Separation of Strong Acid Fraction (SA)
7 Restore, original ether volume to 30
ml, if necessary. Mark flask "NaHCOa"
Shake three times with 15 ml portions »
of 5% NaHCOs and drain aqueous layers
into NaHCOs flask after each shaking.
Acidify aqueous solution in flask by
adding sufficient concentrated HCl
(dropwise because of liberation of CC>2)
until strongly acid to litmus paper.
About 4 ml is sufficient. The solution
becomes cloudy at this point. Set
aside for back-extraction.
Separation of Weak Acid Fraction (WA)
8 Restore original ether volume to 30 ml,
if necessary. Mark flask "NaOH".
Shake three times with 15 ml portions
of 5% NaOH and drain aqueous layers
into "NaOH" flask after each shaking.
Wash once with 15 ml distilled water
and drain this into "NaOH" flask.
CAUTION: Tilt separatory funnel gently
a few times, instead of shak-
ing the first portion, as e-
mulsions often occur.
If a small amount of the water wash re-
mains emulsified, after draining, add
several drops of saturated Na2SO4 so-
lution and shake. This may help to break
the emulsion. Acidify aqueous solution
in flask with concentrated HCL until
strongly acid to litmus paper. About 8
ml sufficient. The solution becomes
cloudy at this point. Set aside for back-
extraction.
Neutral Fraction (N)
9 Mark flask "N". Pour the ether layer
remaining in the separatory funnel in-
to the flask "N". Add about 10 gm of
anhydrous sodium sulfate, cap with
aluminum foil and allow to stand over-
night to dry the ether solution.
B Back Extraction
Strong Acids (SA)
10 Mark flask "SA". Transfer NaHCO3
extract from step 7 to separatory fun-
nel and shake several times, care-
fully, relieving pressure caused by
27-11
-------
Procedures for the Preliminary Separation of Extracts
WEIGHED SAMPLE
add ether, filter
Ether Solution
extract with
Ether Layer
extract with HC1
Ether Layer
extract with NaHCO3
Residue
evaporate, weigh
ETHER INSOLUBLES (El)
Water Layer
evaporate, weigh
I
WATER SOLUBLES (WS)
Water Layer
Make basic, extract
with ether
Water Layer
make acid
extract with ether
Ether Layer
dry, evaporate
and weigh
BASES (B)
Water Layer
Discard
Ether Layer
dry, evaporate
and weigh
Water Layer
Discard
STRONG ACIDS (SA)
Ether Layer
extract with NaOH
Water Layer
make acid
extract with ether
Ether Layer
dry, evaporate
and weigh
Ether Layer
dry, evaporate
and weigh
I
Water Layer
Discard
NEUTRALS (N)
WEAK ACIDS (WA)
27-12
-------
Procedures for the Preliminary Separation of Extracts
evolution of CO2. Shake the NaHCO3
extract, previously made acid, with a
15 ml portion of ether. Caution: Re-
lease CC>2 during first shaking repeated-
ly to avoid pressure build-up.
Drain the aqueous layer into the NaHCC>3
flask. Pour the ether layer into the
"SA" flask. Return aqueous layer to
separatory funnel and repeat, extraction
two more times with 15 ml portions of
ether.
After all the ether layers have been
collected in the "SA" flask, add about
10 gms anhydrous sodium sulfate, cap
with foil and allow to stand overnight
to dry the ether solution. Discard the
aqueous NaHCC>3 portion which should
be almost colorless.
Weak Acids (WA)
11 Mark flask "WA". Shake the NaOH ex-
tract, previously made acid, three
times with 15 ml portions of ether and
collect in flask "WA" as in step 10.
Add sodium sulfate to dry, cap with
foil and allow to stand. Discard aqueous
NaOH portion which should be almost
colorless.
Bases (B)
12 Mark flask "B". Shake the HC1 extract,
previously made basic, three times
with 15 ml portions of ether and col-
lect in flask "B". Add sodium sulfate
to dry, cap with foil and allow to stand.
Discard aqueous HC1 portion.
C Transfer
13 Weigh four flasks and mark them "N",
"WA", "SA", and "B". Record weights
of each. Transfer the corresponding
dried ether solutions from the Na2SC>4
into the weighed flasks by filtering
through filter paper.
14 Evaporate the ether from the above
fractions with the aid of clean, dry cir-
culating air and gentle heat on a steam
bath. The ether solutions should be
removed from the air and heat before
totally dry and allowed to dry spontane-
ously to prevent loss of volatile com-
ponents. Record the weight of the dried
fractions.
D Chromatographic Separation
15 Weigh a 10 ml beaker and record weight.
Dissolve the Neutrals in about 5 ml an-
hydrous ether, transfer most of the Neu-
trals to the beaker and place the remain-
ing few drops in a vial for infrared analy-
sis. Wash the (N) flask with ether and
pour washings into vial also. Dry the
Neutrals contained in the beaker by
spontaneous evaporation and record
weight.
16 Weigh a set of 125 ml Erlenmeyer
flasks and mark them Aliphatics, Aro-
matics and Oxys. Record weights of
each.
17 Fill the chromatographic column with
4 1/2" of silica gel and tap down to
about 4". Place the Aliphatics flask
under the column. Add about 20 ml of
iso-octane to wet the column.
Aliphatics
18 Before the iso-octane reaches the le-
vel of the silica gel, add the Neutrals
which have been previously adsorbed
onto a small amount of silica gel by
stirring in the beaker with a rigid
wire. Elute with 85 ml iso-octane.
Rinse the beaker with a small portion
and add to the column first. Allow
this volume to enter silica gel before
adding the remainder of the iso-octane.
Collect the eluent in the Aliphatics
flask, evaporate to dryness with the
aid of gentle heat and circulating air.
Aromatic s
19 Place the Aromatics flask under the
column. Elute column with 85 ml of
Benzene-rinsing the beaker with a
small portion and adding immediately
as the previous eluent reaches the
level of the silica gel. Allow this vol-
ume to enter silica gel before adding
remainder of benzene. Evaporate
27-13
-------
Procedures for the Preliminary Separation of Extracts
CHROMATOGRAPHIC SEPARATION OF NEUTRALS
NEUTRALS
Adsorb on Silica gel Column
Elute with
Iso-octane
ALIPHATICS
Elute with
Benzene
AROMATICS
Elute with
Chloroform/ Methanol
(1:1)
OXYGENATED COMPOUNDS
with the aid of gentle heat and circu-
lating air. The Aromatics fraction
should be removed from the air and
heat before totally dry and allowed to
dry spontaneously to prevent loss of
volatile components.
20 Place the Oxys flask under the column.
Elute with 85 ml of a 1:1 methanol
chloroform mixture - rinsing the beaker
with a small portion and proceed as
before. Evaporate to dryness with the
aid of gentle heat and circulating air.
21 Record weights of the dried Aliphatics,
Aromatics and Oxys.
NOTE: (All chromatographic solvents
should be redistilled before use)
REFERENCES
1 Shriner, Fuson and Curtin, "The Systematic
Identification of Organic Compounds, "
4th Ed. , John Wiley & Sons, New York,
1956.
2 Cheronis and Entrikin, "Semimicro Qualita-
tive Organic Analysis, " Thomas Y.
Crowell Co. , New York, 1947.
3 Cheronis, "Micro and Semimicro Methods, "
Vol. VI of the series "Techniques of
Organic Chemistry, " Arnold Weissber-
ger, Ed. , Interscience, New York, 1954.
4 Schneider, "Qualitative Organic Microanaly-
sis, " John Wiley & Sons, New York,
1946.
These books deal principally with chemical
methods of identification; Shriner, Fuson
and Curtin and Cheronis and Entrikin are
recommended for those new to the field. For
instrumental analysis and quantitative work,
see standard works in those fields.
27-14
-------
SURFACTANTS AND SYNTHETIC DETERGENTS
R. C. Kroner*
I COMMERCIAL SIGNIFICANCE
Synthetic detergents are a new class of
organic chemicals of extreme commercial
significance. In approximately 20 years sales
have increased from a negligible amount to
about 3. 8 billion pounds per year. They are
of public health significance because almost
every pound of material sold ultimately finds
its way into a sewage treatment plant, a
surface water or a water treatment plant.
o
II
C1nH_.. - O - S - O
12 25 ||
O
Na
2 Cationic, in which the long chain por-
tion of the molecule carries a positive
charge:
A Household users make up about 75% of the
market. Industrial users, which are
rapidly increasing, currently use the
remaining 25%.
B Household uses are well established. In-
dustrial uses include natural and synthetic
fiber processing and dyeing, fur and
leather processing, paper making, brick
and pottery processing, electroplating
and electrocleaning, concrete preparations,
rubber manufacture, formulations as in
soluble oils, insecticides, cosmetics, etc.
II NATURE OF SURFACTANTS(7) (8)
What are surfactants, synthetic detergents?
A Surface active agents are soluble organic
materials which possess the property of
altering the surface or interfacial pro-
perties of their solutions to an unusual
extent when present in low concentrations.
Three types of surface active materials
are presently in use.
Anionic, in which the long chain
^portion of the molecule carries a
negative charge:
CH
C12H25 - ?
CH,,
Cl
3 Nonionic, which does not ionize at all.
C8H17
-O-
C A typical household detergent may contain
the materials shown in Table 1.
D The advantages of synthetic detergents are
stability in low pH waters, no "scums"
with hard waters, superior efficiency,
variety of formulation, cheapness.
E Anionics are used principally in household
cleaners. Cationics are used as sanitizers
in laundering and dish-washing (hospitals,
restaurants, taverns). Nonionics are used
as suds controllers but find principal usage
in industrial applications.
*In Charge, General Laboratory Services, Water Quality Section, DWSPC, SEC, and revised by
Betty Ann Punghorst, Chemist, DWSPC Training Activities, SEC. Reviewed December 1965.
CH. DS.9b. 11.64
28-1
-------
Surfactants and Synthetic Detergents
TABLE 1
CHEMICAL
Surface active agent
Sodium phosphate
Sodium sulfate
Sodium silicate
Methyl cellulose
Fatty amides
Sodium perborate
Perfumes, anti-oxidants
fluorescent dyes, etc.
%
20%
35%
20%
10%
1%
3%
10%
0.1%
FUNCTION
Reduce surface tension
Assist micelle formation,
pH control, metals com-
plexation
Additional electrolyte, builder
Prevents metal corrosion
Prevents redeposition of
soil particles
Foam stabilizer
Bleaching agent
F Cationics and anionics react, forming
precipitates, effectively removing
cationics from solution. Since anionics
are always in excess, cationics pose no
problem in water systems.
G Surface active properties are result of:
64
hydrophobic portion
of molecule
SO Na
o
hydrophilic portion
of molecule
hydrophobic - from Greek, meaning
"water fearing".
hydrophilic - from Greek, meaning
"water loving".
UI PUBLIC HEALTH SIGNIFICANCE
OF SURFACTANTS(7)(8)
Synthetic detergents are said to have adverse
effects on water supplies and treatment
processes.
A Sewage treatment plants have experienced
severe foaming problems, sedimentation
difficulties, decrease in coagulation
efficiencies, changes in biological oxidation
processes.
B Water treatment plants have experienced
coagulation difficulties, foaming and
frothing at plant and in homes, taste
problems, etc.
28-2
-------
Surfactants and Synthetic Detergents
C Natural surface waters may contain in-
creased amounts of phosphates leading
to production of algae growths. Efficiency
of oxygen transfer may be reduced. Foam-
ing may follow heavy rains or washouts.
D A diversity of opinion exists as to the
actual causes of these experiences.
IV ANALYTICAL METHODS FOR
ANIONIC SURFACTANTS
None of the analytical procedures available
are completely satisfactory.
A Methylene Blue
R-SO3Na + methylene blue
—R-SO3Na-MB
The dye complex is soluble in chloro-
form. Remove colored chloroform
layer; read for color.
2 Interferences include NO , NOQ, CNS,
proteins.
,
£t
(2)
B Two Phase Titration
1 Anion + cation -» complex precipitate.
Titrate anion with standard cation; excess
cation reacts with acid dye to form dye
complex in solvent layer.
2 Interferences by soap and protein are
eliminated by pH adjustment. Versene
eliminates Ca and Mg effect.
/ c\
C Infrared Procedure
Sample is treated with activated carbon
which adsorbs organic material. Surfactant
material can be dissolved differentially
with proper solvent. Extract analysis with
infrared will identify the particular sur-
factant and can also be used for quantitative
measurement.
V LABORATORY PROCEDURE: Measure-
ment of Anionic Surfactants by Methylene
Blue
A Reagents
1 Methylene blue solution - dissolve 0. 35
gms methylene blue in one liter of .01
N sulfuric acid.
2 Chloroform, C.P.
3 Sulfuric Acid, 5N
4 Stock ABS Solution, 1.00 mg/ml
5 ABS Working Standard, 0.010 mg/ml
B Glassware
1 Separatory funnels, 250 or 125 ml
2 Volumetric flasks, 50 ml
3 Filtering funnels
4 Pipettes, assorted sizes
C Preparation of Standard Alkyl Benzene
Sulfonate Solutions
1 Weigh an amount of the reference ma-
terial (obtain from Association of
American Soap and Glycerine Producers,
New York) equal to 1. 000 g ABS on a
100 per cent active basis. Transfer
quantitatively to a 1.0 liter volumetric
flask and add 500 ml of distilled water.
Swirl gently until all of the powder is
dissolved, let stand for one half hour
or until most of foam breaks and then
make up to mark with distilled water. Each
ml of this solution contains one mg ABS.
2 Working Standard: Pipette 10.0 ml of
the stock solution into a 1.0 liter volu-
metric flask and make up to mark. Each
ml of this solution contains 0. 010 mg
ABS.
28-3
-------
Surfactants and Synthetic Detergents
D Procedure
1 Add 0, 1.0 2.0,5.0 10.0 20.0 and
25. 0 ml of working standard to separatory
funnels and make up to 100 ml with dis-
tilled water. These standards contain
0, .01, .02, .05, .10, .20 and. 25 mg
of ABS.
2 Add 100 ml of each sample to a separatory
funnel.
3 Add 1. 0 ml of 5N sulfuric acid and 5.0
ml of methylene blue solution to each
sample and standard. Mix well.
4 Add 10 ml of chloroform to each
funnel, invert and shake once a second
for 25 seconds. Allow the chloroform
layer to separate.
5 Draw off the chloroform layer and
filter through a plug of absorbent cotton
into a 50 ml volumetric flask. Repeat
the extraction twice more, using 10 ml
portions of chloroform, collecting the
extracts in the same flask.
6 Rinse the cotton plugs with chloroform
into the 50 ml volumetric flask, make
up to mark with chloroform, mix well
and let stand for 5 to 10 minutes.
Read the optical density of the Reagent
Blank and Standards against chloro-
form as a reference blank at 650 m|i in
a suitable photometer or spectrophoto-
meter and prepare a standard curve.
Read the optical density of each
sample in a similar manner and calculate
mg/1 of ABS in each portion. Report
all results to the second decimal place.
VI SUMMARY
Alkyl benzene sulfonate (ABS), an anionic
surfactant and important constitutent of
syndets, is both a surf ace-active agent and
a biologically "hard" substance. The Public
Health Service in its Revised Drinking Water
Standards of 1962 has stated that ABS should
not be present in excess of 0. 5 mg/ 1 in any
water supply where a more suitable supply
is available. Various methods have been
formulated for the detection of anionic sur-
factants, the most popular one being the
methylene blue extraction procedure.
28-4
-------
Surfactants and Synthetic Detergents
REFERENCES
1 Bibliography on Synthetic Detergents in
Water and Wastes. Public Health
Service. Robert A. Taft Sanitary
Engineering Center (Basic and Applied
Sciences Branch, Division of Water
Supply and Pollution Control) . June,
1964.
2 Edwards, Gail P., and Glnn, Martin E.
Determination of Synthetic Detergents
in Sewage. Sewage and Industrial
Wastes. 26: 945. 1954.
3 Hill, W. H., Shapiro, M. A., and Kobayashi,
Y. Determination of Alkyl Benzene Sul-
fonate in Water. Jour. Amer. Water
Works Assoc. 54: 409-416. 1962.
5 Longwell J., and Maniece, W. D. Deter-
mination of Anionic Detergents in
Sewage, Sewage Effluents and River
Waters. Analyst. 80: 167. 1955.
6 Sallee, E.M., et al. Determination of
Trace Amounts of Alkyl Benzene Sul-
fonates in Water. Anal. Chem. 28:
1822. 1956.
7 Sawyer, Clair N., and Ryckman, Devere
W. Anionic Synthetic Detergents and
Water Supply Problems. Jour. Amer.
Water Works Assoc. 49: 480-490. 19~57.
8 Task Group Report. Characteristics and
Effects of Synthetic Detergents. Jour.
Amer. Water Works Assoc. 46: 751-
774. 1954.
Jones, J. H. General Colorimetric
Method for Determination of Small
Quantities of Sulfonated or Sulfated Sur-
face Active Compounds. Jour, of
Associated Official Agri. Chemist. 28:
3987 1945.
28-5
-------
INFRARED INSTRUMENTATION
D. G. Ballinger*
I INTRODUCTION
In infrared work the spectroscopist is using
a spectral region which is not visible to the
human eye. The types of visual comparators
commonly used in the laboratory cannot be
applied to measurements in the IR. Thus the
analyst is completely dependent upon an elec-
trical system for his evaluations.
Because of the specific nature of the IR
spectra of most substances, the full record-
ed spectrum, with its characteristic peaks
is generally used. The complexity of the IR
spectrum rules out the possibility of manual
plotting of absorption curves. Sensitive re-
corders, having rapid response,are required.
The region generally examined extends from
2.0 - 15(1 (Fig. 1). Some research instru-
ments are capable of scanning a much wider
range, from 1.0 - 40^. In this portion of
the spectrum energy levels are low and strong
absorption bands are found.
As in all analytical spectroscopy, the measure-
ment system involves a source of radiant
energy, an arrangement for placing the sam-
ple in the incident beam, and a detector or
receptor for measuring the changes in energy
caused by absorption.
II SOURCE OF RADIANT ENERGY
A Nernst Glower
This type of element consists of a mix-
ture of rare earth oxides bonded into a
small rod. When electrically heated this
source gives off infrared energy of the
desired wavelengths.
B Globar
In many newer instruments a rod of bond-
ed silicon carbide is used. Like the
glower, the energy emitted is in the de-
sired region of the spectrum.
-2 ° ° 2 ° 2
10 A 1 A 10 A In 10% 1 cm
!
f>
\
<
<
GAMMA
RAYS
X-RAYS
ULTRA
VIOLET
VISIBLE
« MICRO- AND
RADIO WAVES
INFRARED
Figure 1 - ELECTROMAGNETIC SPECTRUM
'-Supervisory Chemist, Technical Advisory and Investigations Section, DWS&PC, SEC,
Reviewed December 1965.
CH. MET. 13. 12.63
29-1
-------
Infrared Instrumentation
C Nichrome Wire
A few IR instruments use an electrically
heated nichrome wire as a source.
D Monochromator
In order to provide a spectrum for study
the IR instrument must disperse the energy
from the source into small increments and
selectively direct these wavelength com-
ponents through the sample. These functions
are accomplished by the monochromator.
Since the glass optics used in visual range
instruments will not transmit infrared
energy, a prism of another material must
be used. (Fig. 2) shows the optical ma-
terials commonly used, with the effective
range of each.
In order to maintain constant energy level
with changing wavelength, the slit is
"programmed". That is, the slit changes
as the wavelength increases, compensating
for the loss of energy at the longer wave-
lengths.
Ill CELL ASSEMBLY AND ,
CHARACTERISTICS
A Geometry
In most photometric instruments the
radiant energy reaches the sample after
being dispersed into selected wavelengths.
In IR spectrophotometers it is possible to
place the sample in the undispersed beam
and then pass the beam through a suitable
monochromator to the detector. In the
double beam instrument shown in Pig. 3
the energy from the source is split into
two equal beams. One beam passes
through the sample.the other is unchanged
and thus becomes a reference. The re-
corder indicates the difference in energy
between the two beams; that difference
is the result of absorption of radiant energy
by the sample.
B Cell Characteristics
1 Materials
The factors which govern the selection
of optical materials in the monochromator
MICRONS
10 , 15
20
25
GLASS
.. QUARTZ
LiF
CaFr
NaCl
Figure 2 - OPTICAL MATERIALS
29-2
-------
Infrared Instrumentation
Figure 3 - I-R SPECTROPHOTOMETER
DETECTOR
MONO-
CHROMATOR
SOURCE
SAMPLE
also apply to cell materials. See j ig.
-------
Infrared Instrumentation
filled by means of a hypodermic syringe.
After filling, the ports are sealed with
Telfon plugs.
C Special Cells
1 Smears
When non-volatile materials are being
examined, the sample can be smeared
directly onto a cell window, with or
without a covering window.
2 KBr Technique
Crystalline solids cannot be examined
directly, due to light scattering by the
crystals. A technique has been develop-
ed using potassium bromide, whereby a
solid can be mixed with KBr and com-
pressed into a very thin sheet. Since
the KBr is transparent in the IR, these
discs permit the examination of the
solid sample. Preparation of the KBr
discs is difficult but opens the way to
the examination of a wider range of
organic and inorganic materials.
3 Gas Cells
Because of the high sensitivity of the
IR Spectrophotometer, it is possible to
obtain the spectrum of gases as well as
liquids and solids. Obviously the mole-
cular density of gases necessitates a
much longer light path. Special gas
cells have been developed with path
lengths from 5 cm to 10 meters. The
5 cm cells have a single pass light
path, while the longer paths are obtained
by multiple passes, as shown in Fig. 5.
IV DETECTOR «
A Theoretical
Radiant energy in the IR region is actually
heat radiation. The absorption of energy
at various wavelengths is the absorption
of heat. Thus an infrared detector is a
heat-measuring device. The most com-
monly used detectors are the thermocouple
and the Bolometer. Although these measure
the ambient temperature as well as the
signal, it is only necessary to measure the
difference between the sample beam and
the reference beam. Further, since ra-
diant energy is being measured, small
temperature differences in the sample
have no effect.
B Thermocouple
The bimetallic junction of the thermo-
couple is capable of measuring energy
changes over the whole IR range. Because
a very rapid response is required, special
thermocouples have been designed for IR
applications.
C Bolometer
Certain metals have the property of high
thermal resistivity. That is, the electri-
cal resistance of the metal changes greatly
with change in temperature. The bolo-
meter consists of an element of such a
metal mounted as a target in the IR beam.
Changes in resistance, produced by energy
flucuations, are converted to electrical
impulses and fed to the amplifier circuit.
29-
-------
Infrared Instrumentation
Figure 5 -ONE METER GAS ABSORPTION CELL
(Courtesy Perkin - Elmer Corporation)
29-5
-------
INFRARED IDENTIFICATION OF ORGANIC COMPOUNDS
4000 3000
100
2000
1500
CM I
1000 900
BOO
700
•1 V 10 II
WAVEICNOTH (MICRONS1
I/
1 t 14
15
SPECTRUM NO. //
SAMPLE
ORIGIN ~1
PURITY ^J
PHASE
THICKNESS^A'^'V. *
LEGEND
1
•)
DATE •*:/•> •••*?._ ..
OPERATOR ^'^ _.
REMARKS
0-CH^CH;,
in
t>
|
THC PERKIN-EIMER CORPORATION, NORWALK, CONN.
CH. MET. 19. 12.63
-------
Infrared Identification
7 "^ 8 9 10
WAVELENGTH (MICRONS)
SPECTRUM NO. 9
SAMPLE
M'rjogei/zf^f
ORIGIM
PURITY *£•*
PHASE 't'f'o
THICKNESS c if*»
LEGEND
1
2
DATE ~- •f-J-f
OPERATOR &J0
REMARKS
r-
-\
1 )
V
1
o
4000 3000
100
2000
1500
THC PERKIN ELMFR COPPORATION, NORWAIK, CONN.
CM-' 1000 900 800 700
3 4 "5 67 8 9 10
WAVELENGTH (MICRONS)
_ I L I wf,ot)is>0
THICKNESS 02 ***___
LEGEND
1
•).
DATE ^-/^-J'-f _
OPERATOR ^& . _
REMARKS
ft*"
(^S-rS
0
V
,
>
s,
'
THE PERKIN-ELMER CORPORATION, NORWAIK, CONN.
29-7
-------
Infrared Identification
4000 3000
1'4AJ,'yk'.J, l-lri - 1—I -
2000
1500
CM'
1000 900 800-
?00
WAVELfcNGTH (MICRONS)
SPECTRUM NO._£__
SAMPLE . .
ORIGIN.
MJRITY_. '_! _
LEGEND
DATE •?--v.
LEGEND __ __. -
2,
DATE 3/zz/Jy. _ __
OPERATOR #~&
REMARKS
'£
Z
o
THE PERKIN-EIMER CORPORATION, NORWAIK, CONN.
-------
Infrared Identification
4000 3000
loo1"1"""''1"'
2000
1500
CM-'
1000 900
800
700
7 8 9 10 11
WAVELENGTH (MICRONS)
SPECTRUM NO. j>
SAMPLE
dctrJirf
ORIGIN
PURITY ^
PHASE <7-
COM
• -a
O
4000 3000
2000
1500
THE PERKIN.EIMER CORPORATION, NORWALK, CONN.
CM-' 1000 900 800 700
8 9 10 11 12 13 14 15
WAVELENGTH (MICRONS)
SPECTRUM NO._JI
SAMPLE
ORIGIK
PURITY -"V
PHASE ^rjL
THICKNESS ^«*
LEGEND.
1
2
DATE f't ->?f . _
OPERATOR c:f<3. -
REMARKS^.
•»«,*« >£*?«*•,«
/-<•-<.#,-! ",- '/I'-. ^
0--. u, < H. . tt, -i »,
7
SS
o
THE PERKIN.EIMER CORPORATION, NORWALK, CONN.
29-9
-------
Infrared Identification
CM' 1000 900 800
700
.;:L
WAVELENGTH (MICRONS)
SPECTRUM NO. ?
SAMPLE
ORIGIN .
PURITY ACT
PHASE *'(>•"*
THICKNESS 0* w
If GtND. ...
1
7
DATE 4-a-j-r
REMARKS .. _.
we*-**
' V
'c
5
Z
O
THE PERKIN-EIMER CORPORATION, NORWAIK, CONN
4000 3000 2000
1500
CM-'
1000 900 800
.-i-t-J j-4_i_J_L-t J—L-L-J-l-^-
5 6 7 8 9 10
WAVELENGTH (MICRONS)
II 12 13 14
SPECTRUM NO._£j?
SAMPLE (4)
ORIGIN
PURITY Jcj
PHASE •
THICKNESS 6^/i^cv
LEGEND
1.
2
DATE ^-A-/?_
OPERATOR -&«^
REMARKS
AU-ci'-^i. -*- t-X' -:>*•
^"
f^2
Z
O
CH.OTS.lab.J.lj.S?
THE rCRKIN-ElMER CORPORATION, NORWAIK, CONN
20-10
-------
Infrared Identification
2000
1500
CM-'
1000 900
800
700
7 n 9 10
WAVELENGTH (MICRONS)
12 13 U 55
SPECTRUM NO._^:
SAMPLE
ORIGIN
PURITY <*."
PHASE &£*•
THICKNESS.,!/****- .
LEGEND,
1.
2.
DATE J-//-4-?
OPERATOR ^>"C -
LEGEND .
1
2
DATE a - //-s.?
OPERATOR P4£
REMARKS
O
CH.UrS.lob.l.li.SV
THE PERKIN-EIMCR CORPORATION, NORWALK, CONN.
29-11
-------
Infrared Identification
4000 3000
rooo 900
800
700
4567 9 10 11 V,
WAVELENGTH (MICRONS)
SAMPLF NO, 13
SOURCE pl^u ^"'fe "'-
LOCATION
DATE SAMPLED- . .. ...
TYPp oMipulir."
REMARKS
PHASP '-'vi'"^*
OPFRATOR <*?
DAT El v^-i^
*
.,3
|
r t/i w
o o >
O C £
J> a> TI
H o r-
o m m
z r
o
4000 3000
I . . . .
800
700
7 8 9 10
WAVEL£NGTH (MICRONS)
CAMPLE NO H
SOURCE W-MET^VL wJiLik/e.
LOCATION
DATE SAMPLED
TYPp JWJIWICH-
REMARKS _
-
PHASE L|1UI-CL
OPERATOR "w
DATE ^M-J .
A-^'**
y XH-
i- u> u>
o o >
o c C
«> a) -o
H o I-
o m m
z
29-12
-------
Infrared Identification
4000 3000
CM-1
1000 900
800
700
WAVELENGTH (MICRONS)
9AMP| f NO IS'
SOURCE PURE BEWZVL ETOE«
LOCATION
DATp ^AMPI FT}
REMARKS
PHA«1F VW'C
DATE VC-J.3
v^^-ft
O
r>
5
z
Jl 4
?!
a •
n i
f1! r
i
4000 3000
100
2000
1500
CM-i
1000 900
800
700
U
60
i40
20
INFUACCMP ,3K^ 137.1281
7 8 9 10 11 12 13 14 15
WAVELENGTH (MICRONS)
SPECTRUM NO. Ifc
SAMPLEJ^f1^^1-^ *CiO
—
ORIGIN
PURITY fyhE __
PHASE LIQUID
THICKNESS_iJiy[rL__
LEGEND
1
2.
DATE a-l'-«-J
OPERATOR__S^ __
REMARKS
tHjCH^CHj^NjCH^CMjCHtCt^
in
£
s
Tl
THE PERKIN-ELMER CORPORATION, NORWAUC, CONN.
29-13
-------
Infrared Identification
DATA SHEET - ORGANIC LABORATORY
SOLUBILITY SEPARATION
WEIGHTS
ORIGINAL SAMPLE
GM
BASES
GM
STRONG ACIDS
GM
WEAK ACIDS
GM
NEUTRALS
GM
REMARKS
CHROMATOGRAPHIC SEPARATION
OF NEUTRALS
WEIGHTS
ALIQUOT USED
GM
ALIPHATICS
GM
AROMATICS
GM
OXYS
GM
REMARKS
SUMMARY OF DATA
BASES
STRONG ACIDS
WEAK ACIDS
NEUTRALS
TOTAL RECOVERED
% RECOVERY
GM
GM
%
29-14
-------
Infrared Identification
N. B. COLTHUP
29-15
-------
Infrared Identification
SPECTRA-STRUCTURE CORRELATIONS
==*pMl=l|i
j- faj:jtfe=±=t:
fc^fei;:E:^lfe:
?
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n
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5
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c
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IE S"e •
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•^ ^1
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nl 3
Jd O
0 ^
29-16
-------
CHEMICAL OXYGEN DEMAND AND COD/BOD RELATIONSHIPS
R. J. Lishka*
I DEFINITION
Chemical Oxygen Demand is an analytical
parameter of pollution which measures the
chemically oxidizable material in a liquid
sample.
A A variety of terms have been used to
describe this parameter.
1 OC - oxygen consumed
2 COD - chemical oxygen demand
3 - complete oxygen demand
4 - dichromate oxygen demand
B The COD test may be compared to other
similar tests that attempt to characterize
materials by "shotgun" procedures.
1 "iodine number" - for evaluating fatty
acids
2 "substances decolorizing perman-
ganate" - used for assaying reagent
grade chemicals
3 "chlorine demand" - for evaluating
oxidizable materials in water
II A chemical oxidation demand test is
needed for many reasons, chief among
which are:
A Chemical oxidation is comparatively
rapid as contrasted to biological
oxidation.
B Chemical oxidation is not subject to as
many variables (seeding, toxicity, etc.)
as is a biological system.
C The chemical procedure requires less
equipment and is less expensive to run.
D In the presence of high toxicity, the
chemical procedure may be the only way
to determine the organic load.
Ill The concept of the COD is almost as old
as the BOD. Many oxidants and variations
in procedure have been proposed, but none
have been completely satisfactory.
A Ceric sulfate has been investigated, but
in general it is not a strong oxidant.
B Potassium permanganate was one of the
earliest oxidants proposed and until re-
cently appeared in Standard Methods (9th
ed.) as a standard procedure. The re-
sults obtained with permanganate were
dependent upon concentration of reagent,
time of oxidation, temperature, etc. , so
that results were not reproducible.
C Potassium iodate or iodic acid is an
excellent oxidant but methods employing
this reaction are time-consuming and
require very close control.
D A number of investigators have used
potassium dichromate under a variety of
conditions. The method proposed by
Moore at SEC is now accepted as the
standard procedure because of the
reproducibility, ease of manipulation,
applicability to a wide variety of samples,
and the use of a stable, primary standard
reagent. *
IV The present method for chemical oxygen
demand has been thoroughly studied and
statistically evaluated by replicate compari-
son with other methods.
A The procedure consists of refluxing a
sample containing organic material with
sulfuric acid and an excess of standard-
ized potassium dichromate. During the
*Chemist, Analytical Reference Service, Training Program and revised by J. W. Mandia, Chemist
DWS&PC Training Activities. Reviewed December 1965.
CH. O. oc. 5d. 8. 65
30-1
-------
Chemical Oxygen Demand and COD/BOD Relationships
reflux period the chemically oxidizable
organic material reduces a Stoichio-
metric equivalent amount of dichromate,
the remainder of which is measured by
titration with standard ferrous ammonium
sulfate. The amount of dichromate re-
duced (amount of dichromate added -
amount of dichromate remaining =
amount reduced) is a measure of the
amount of organic material oxidized.
B Factors to be observed in the proper con-
trol of the reaction are as follows:
1 The potassium dichromate and ferrous
ammonium sulfate reagents must be
standardized.
2 The volume of dichromate solution
used must be accurately measured
(25.0 ml).
3 The volume of sample must be accur-
ately measured,
4 The concentration of sulfuric acid in
the refluxing solution should be 50% by
volume.
5 Sufficient time should be allowed to
secure complete oxidation of sample,
usually not more than two hours.
V As in the case of biological oxidation, not
all organic compounds are oxidizable by wet
chemical methods.
A Some compounds are completely oxidizable
with no difficulty. Common among these
are sugars, branched chain aliphatics,
and substituted benzene rings.
B Other compounds are not oxidized by this
method, chief among which are benzene,
pyridine, and toluene.
C Some compounds are only partially
oxidized, but the oxidation efficiency
may be increased by the use of silver
sulfate as a catalyst in the refluxing
solution. Straight chain acids, alcohols,
and ammo acids are examples of compounds
which can be completely oxidized with the*
assistance of the silver sulfate catalyst.
VI The COD test(1) measures not only the
chemically oxidizable organic material but
also any chemically oxidizable inorganic
material (e. g. , chloride ion) contained in
the sample. Chloride ion at a concentration
of 200 mg/1 or more (10 mg Cl/50 ml sample)
represents a positive interference in this
procedure.
A By actual measurement, the correction is
(mg/1 C1X 0.23 = mg/1 COD). This
correction is not applied when the silver
sulfate catalyst is used because of pre-
cipitation of the silver as AgCl.
B It has been established that chlorides are
not always quantitatively oxidized in the
presence of organic matter. In the pre-
sence of high concentrations of ammonia,
organic amine, or nitrogenous material,
a continuous reduction of dichromate
occurs. The mechanism for this reduction
is thought to involve a series of cyclic
changes from chlorine to chloride through
the formation of chloramines. The addi-
tion of silver sulfate at the start of the
digestion can lead to error because of the
unknown degree of chloride oxidation. In
addition, much of the catalytic action is
lost due to precipitation of the silver ion.
Furthermore, in a sample containing a
high concentration of chlorides, the
application of the chloride correction may
result in a negative value for the COD.
C To overcome the difficulties presented by
chlorides, it was found that chloride could
be eliminated from reaction by a complex-
ing technique using mercuric sulfate. *1)
By the simple expedient of adding both
sulfate and silver sulfate to the sample,
interference from chloride is avoided,
and the catalytic action of the silver sul-
fate is retained.
D COD Method for Highly Saline Waters
The maximum COD that can be measured
using a 50 ml sample and 25 ml 0. 25 N
dichromate is 1,000 mg/1.
30-2
-------
Chemical Oxygen Demand and COD/BOD Relationships
1 A sample containing 10 g/1 chloride
(50% sea water) has a COD of 2, 300
mg/1 due to chloride.
2 When Ag2SO4 is used as a catalyst,
chloride first must be removed.
Ag2S04 + 2C1" ix 2,AgCl,
AgCl is not completely oxidized during
the test. A correction cannot be made
for the AgCl which has not been oxidized.
3 The dissociation constant of AgCl in
distilled water is 1. 56 X 10'10".
4 Hg++ in HgSO4 forms a chloride with
a dissociation constant at 25°C of
2. 6 X 10"-^ The greater reduction
of chloride COD occurs in the absence
of Ag2SO4 catalyst.
5 A correction factor of 0. 014 X mg/1
Cl" is applied for a 2 hour COD
digestion using 10 mg HgSO4/mg
chloride in the COD mixture in the
presence of lgAgSO4-
'II It is possible to calculate the theoretical
chemical oxygen demand for organic com-
pounds if the oxidation reaction is known.
A Sample calculation for glucose;
B Sample calculation for phenol:
1. 000 g X
C6H5OH + 7 O2—>• 6CO2 + 3 H2O
94 g 224 g
1.000 X 224 _ 2 383
Oxygen
Theoretical COD =
94
1.000 g X
C6H12°6
180 g (6 X 32) g
C°
H2°
X - — -
Where
180
32
1. 000
X
molecular weight of glucose
molecular weight of oxygen
experimental weight (in g) of
glucose
weight of oxygen required to
oxidize 1. 000 g of glucose
VIII It is not possible to establish fixed
relationships between BOD and COD measure-
ment until a particular sample has been
characterized by both parameters.
A If the sample is primarily composed of
compounds that are oxidized by both
procedures (BOD and COD) a relation-
ship may be established.
1 The COD procedure may be substituted
(with proper qualifications) for BOD.
2 The COD may be used as an indication
of the dilution required for setting up
BOD analysis.
B If the sample is characterized by a pre-
dominance of material that can be chemi-
cally oxidized, but not biochemically,
the COD will be greater than the BOD.
Textile wastes, paper mill wastes, raw
sewage, containing high concentrations
of cellulose have a high COD, low BOD.
C If the situation as in item VIII B (above)
is reversed, the BOD will be higher than
the COD. Distillery wastes or refinery
wastes may have a high BOD, low COD.
IX The present standard method for COD is
commonly used for characterizing samples
of high organic content. However, the
method with some alteration may be used for
analysis of samples containing low organic
content, i. e. , from 5 to 50 mg/1 COD.
A The normality of potassium dichromate
and ferrous ammonium sulfate is reduced
from 0. 25 N to 0. 025 N.
30-3
-------
Chemical Oxygen Demand and COD/BOD Relationships
B Special precautions are taken to insure
replicable results:
1 Condensers are plugged with glass
wool to eliminate airborne dust.
2 Distilled water used for dilution must
be of extremely good quality.
3 More indicator must be used to obtain
clean end point.
4 High quality sulfuric acid, with no ^^
appreciable demand, is required. "^ <\
X Using the revised COD procedure for
stream samples, estimates of the probable
BOD ratio at a particular stream point may
be made from a series of previous repre-
sentative COD/BOD ratios.
A In stream surveys, the COD/BOD ratios
from representative sampling points
give information on general stream condi-
tions, location of pollution, ability of the
stream to oxidize the waste load and
relative degree of biological stability.
XI The low-level chemical oxygen demand
test can only be used for samples having a
COD < 100 mg/1. For stronger wastes, the
standard test must be used.
REFERENCES
1 Dobbs, Richard A. , and Williams,
Robert T. Elimination of Chloride
Interference in the Chemical Oxygen
Demand Test. Anal. Chem. 35:1064.
/'" 1963.
2 Standard Methods for the Examination of
Water and Wastewater. llth Edition.
American Public Health Association,
American Water Works Association,
Water Pollution Control Federation,
1960.
3 Cripps, J. M. and Jenkins, D. A COD
Method Suitable for the Analysis of
Highly Saline Waters, JWPCF 36,
1240-1246. 1964.
30-4
-------
LABORATORY FOR CHEMICAL OXYGEN DEMAND DETERMINATION
R.J. Lishka*
I REAGENTS AND EQUIPMENT
A Potassium Dichromate Solution - 0. 25N
B Ferrous Ammonium Sulfate Solution 0. 25N,
Approximately
C Sulfuric Acid - Concentrated
D Mercuric Sulfate - Analytical Reagent
E Silver Sulfate - Analytical Reagent
F o-Phenanthroline Ferrous Indicator
(Ferroin)
G Flasks, Erlenmeyer, 500 ml, 24/40
E Joint
H Condenser, Friedrichs Reflux 24/40
IB Joint
I Burette, 50 ml
J Glass Beads or Porcelain Chips
II PROCEDURE
A Measure 50 ml of sample or aliquot diluted
to 50 ml with distilled water, and place
in 500 ml Erlenmeyer flask then add:
1 1 gram mercuric sulfate
2 5 ml concentrated H^SO - swirl to
dissolve mercuric salt.
3 25 ml 0.25 N
4 70 ml concentrated H^SO (Cautiously)
5 0. 75 gram Ag SO
6 Several glass beads or porcelain chips
B Mix well by swirling flask.
C Connect flask to condenser and reflux for
two hours.
D Wash down the condenser with distilled
water and cool to room temperature.
E Add 10 drops of o- phenanthroline ferrous
indicator and titrate to a red end point
with standardized ferrous ammonium
sulfate solution.
F Carry a blank, consisting of 50 ml of
distilled water, through the same
procedure.
G Standardization of ferrous ammonium
sulfate solution
1 Pipette 25 ml of 0. 25 N K Cr O into
a 500 ml Erlenmeyer Mask.
2 Add 250 ml of distilled water.
3 Add 50 ml cone. H SO
£ ri
4 Add 10 drops of o-phenanthroline ferrous
indicator and titrate to a red end point.
5 Calculation
Normality of Ferrous soln. =
25 X 0.25
ml ferrous soln.
Ill CALCULATION OF COD
COD mg/1 =
a - b X normality of ferrous soln.
X 8000
ml of sample
a = ml ferrous ammonium sulfate used
for blank
b = ml ferrous ammonium sulfate used
for sample
*Chemist, Analytical Reference Service, Training Program, SEC. Reviewed December 19G5.
CH. O. oc.lab. 1. 12.64 30-5
-------
Lab. for COD Determination
IV SPECIAL DIRECTIONS FOR
LOW-LEVEL COD
A Use 0. 025 N potassium dichromate
solution and 0.025 N ferrous ammonium
sulfate solution instead of the 0.25 N
reagents. The procedure and calculation
are unchanged.
B The ferrous ammonium sulfate solution
is not stable and must be standardized
daily.
C Keep the reflux apparatus assembled
when not in use.
D The outlet tube of the condenser should
always be lightly plugged with glass
wool, both during storage and when in use.
E Before disconnecting the flask, wipe the
condenser and the flask neck with a damp
cloth to remove dust particles.
F Periodically, the glass apparatus should
be steamed out to remove trace organic
contamination, using the following
procedure:
1 Add 50 ml distilled water to the flask.
2 Carefully add 50 ml cone, sulfuric
acid and mix thoroughly.
3 Connect the flask to the condenser but
do not turn on the water supply.
4 Apply heat to the flask until the acid
mixture boils and steam emerges from
the condenser.
5 Remove heat, cool, and discard the
acid mixture.
REFERENCES
1 Dobbs, Richard A., and Williams,
Robert T. Elimination of Chloride
Interference in the Chemical Oxygen
Demand Test. Anal. Chem. 35; 1064
1963.
2 Standard Method for the Examination of
Water and Waste water, llth Edition.
APHA. AWWA. WPCF. 1960.
30-6
-------
INTRODUCTION TO GAS-LIQUID CHROMATOGRAPHY
Betty Ann Punghorst*
I INTRODUCTION
A Definition
Gas-liquid chromatography is an analytical
method for the separation and identification
of a mixture of volatile (usually organic)
components in a sample. As with any
chromatographic technique the column
consists of two phases, the immobile or
stationary phase (a liquid on an inert solid
support), and the mobile phase (an inert
gas). The column functions to separate
the sample components because they have
varying vapor pressures and affinities for
the stationary phase. In many ways the
column behavior resembles that of
fractional distillation. The partition which
occurs between the mobile and immobile
phases will thus cause the components to
proceed through the column at varying
rates. The separation is recorded and
quantitated by the detector system.
B Advantages
1 GLC can be used to separate compounds
of similar boiling points which cannot
easily be separated by distillation. (See
Table 1)
2 GLC can be extremely sensitive; for
example, using the electron capture
detector it is possible to "see" pico-
gram (10~12) quantities.
Table 1. SEPARATIONS BY GLC
Compounds
3- Methylcyclohexene
(B.P. 104°C) and
4-Methylcyclohexane
(B.P. 103°C)
Cyclohexane (B. P.
80. 8°C) and Benzene
(B.P. 80.2°C)
Reference
Aerograph Research
Notes (Spring 1964)
Chromosorbe News-
letter (FF-104)
C Disadvantages
1 Due to the extreme sensitivity possible
it is often necessary to apply extensive
clean-up techniques.
2 The many variables of the technique
require a skilled analyst.
II COMPONENTS OF A GAS
CHROMATOGRAPH (See Figure 1)
A Gas Supply
The mobile phase (carrier gas) trans-
ports the sample components through the
column into the detector. The type of
gas used varies with the detector (See
Table 2.)
B Injector
Liquid samples are manually introduced
into the heated injector block through a
rubber septum by means of a syringe.
Automatic liquid injectors as well as in-
jection systems for solid and gaseous
samples are commercially available.
C Column
The vaporized sample enters the column
which can be glass or metal and of varying
length (I1 - 20') and diameter (1/8" - 1/4").
The column is packed with the stationary
(immobile) phase and contained within a
constant temperature oven.
1 Solid support
The solid support should have a large
surface area yet be inert so that active
sites will not cause adsorption of sample
components. Diatomaceous earths,
teflon and glass beads have been used.
(See Table 3.)
*Chemist, DWSPC Training Activities, SEC.
CH. MET.cr.5. 12.65
31-1
-------
Introduction to Gas-Liquid Chromatography
PRESSURE
REGULATOR
AUTOMATIC STRIP
CHART RECORDE
0S1LICOME RUBBER
STOPPERS
®1/16" DIAMETER
S.S.TUBING
Figure 1. COMPONENTS OF A GAS CHROMATOGRAPIT
Table 2. CARRIER GASES
Detector
Thermal conductivity
Microcoulometric
Flame ionization
Electron capture
Carrier gas
Helium (Purified,
Grade A)
Helium (Purified,
Grade A)
Hydrogen (Purified)
Nitrogen or a mixture
of 95% argon and 5%
methane (Purified)
Table 3. SOLID SUPPORTS
Support
Chromosorb P
(Diatomaceous Earth)
Chromosorb W
(Diatomaceous Earth)
Chromosorb G
(Diatomaceous Earth)
Chromosorb T
(Teflon)
4.8
1.2
0.5
7.0-8.0
2 Stationary liquid
The separation and partition occurring
in the column is directly affected by the
choice of stationary liquid. For ex-
ample, in the separation of benzene
(B. P. 80. 1°C) and cyclohexane (B. P.
80. 8°C), the choice of a non-polar phasi
such as hexadecane results in benzene
preceding cyclohexane off the column.
However, if a more polar phase such as
benzylbiphenyl is chosen cyclohexane
precedes benzene. Table 4 shows some
typical stationary liquids and their uses
(NOTE: One requirement for any liquid
is that it have a high boiling point so tha
it will not boil off the column)
Surface area (m2/gm) D Detector
The detector or brain of the gas chroma-
tograph senses and measures the quantity
of sample component coming off the columr
The detector should be maintained at a
temperature higher than the column so that
condensation does not occur in the detector
block. Several types of detectors are in
use today.
*Reproduced (with permission) from Chemist
(37:11, p. 13. November 1964).
31-2
-------
Introduction to Gas-Liquid Chromatography
Table 4. STATIONARY LIQUIDS
Stationary liquid
Used to separate
Silicone Oils QF-1, Dow Corning
200, and Dow 11
Silicone Oil SE-30
Benzyl-Cyanide-Silver Nitrate
Polyethylene Glycol
Cyano Silicone
Chlorinated hydrocarbons
pesticides
Homologous series of n~ alkanes
Homologous series of olefins
Amines
Steroids
1 Thermal conductivity
This detector consists of a Wheatstone
bridge two arms of which are thermal
conductivity cells each containing a
small heated element. When only carrier
gas is flowing through both the sample
cell and reference cell, the resistance
of the heated element is constant in both
cells. The bridge remains balanced and
baseline is recorded. However, when
carrier gas plus sample component enter
the sample cell, the thermal conductivity
in that cell changes thus also producing
a change in the resistance of the heated
element. The bridge becomes unbalanced
and a peak is recorded. The main dis-
advantage of the TC cell in water pollution
work is its lack of sensitivity.
2 lonization detectors
a Flame
This detector consists of a flame
situated between a cathode and anode.
As carrier gas alone burns, some
electrons and negative ions are pro-
duced which are collected at the
anode and recorded as baseline.
When carrier gas plus sample com-
ponent are burned, more electrons
and negative ions are produced which
result in a peak on the recorder.
The detector is capable of "seeing"
nanogram quantities of organic com-
pounds; however, the detector is
sensitive to all organic compounds.
This lack of specificity produces dis-
advantages in the analysis of water
extracts which contain a variety of
naturally occurring organics.
b Electron capture (See Figure 2)
This detector consists of a radiation
source (e.g., tritium) capable of
producing slow electrons in a carrier
gas such as nitrogen. The electrons
collected at the anode are recorded
as baseline. When sample com-
ponents which have an electron
affinity (e.g., chlorinated hydro-
carbons) enter the detector, electrons
are "capture". The subsequent de-
crease in current is recorded as a
peak. The detector has the advantage
that it is extremely sensitive (pico-
gram range) and is somewhat selective.
c Thermionic
A recent adaptation of the flame
ionization detector shows promise
for the specific analysis of organic
phosphorus compounds. The cathode
of the conventional flame ionization
detector is coated with sodium salt.
When compounds containing phos-
phorus emerge from the column, they
give 600X the response with this de-
tector as with the conventional flame.
3 Microcoulometric
This highly specific detector consists
of titration cells for the measurement
of chloride-containing and sulfur-
containing compounds. The sample
component emerging from the column
31-3
-------
Introduction to Gas-Liquid Chromatography
GAS EXHAUST
RADIOACTI VE
TRITIUM
FOIL
—-tt-ANODE
TO
ELECTROMETER
ELECTROMETER
COLUMN
LOSSOFe"
REDUCES CURRENT
Figure 2. ELECTRON CAPTURE DETECTOR (Wilkens Instrument Co.)
is combusted to produce HC1 or SO2>
respectively. HC1 is continuously titrat-
ed by silver ions present in the cell; the
amount of current required to regenerate
these silver ions is recorded as a peak.
The system for sulfur containing com-
pounds is analogous except that SO2 pro-
duced is continuously titrated by 1% which
is subsequently regenerated. Although
less sensitive (by approximately a factor
of 10) than electron capture, this de-
tector is finding wide use in pesticide
analysis.
E Recorder
The recorder system registers the response
of the detector to sample components. In
the case of ionization detectors, it is often
necessary to employ an electrometer in
order to amplify the small current changes.
Expensive integration and digital read-out
equipment is also available to facilitate
measurement of peak areas.
Ill QUALITATIVE ANALYSIS
A Retention Time
The retention time of a sample component
is defined as the time it takes for that
component to travel through the column.
There are a number of variables which
affect the retention time of a compound.
1 Physical parameters of column
operation
a Column length
b Column temperature
c Carrier gas flow rate
2 The nature and amount of stationary
liquid itself.
For a given set of column conditions, a
specific compound will have a specific
retention time (see Figure 3 and Table 5)
Various column and detector combination,
can be used to confirm identification.
B Retention Volume
Retention volume is defined as the total
volume of gas required to move a com-
ponent through the column.
RETENTION
VOLUME (RT
= RETENTION X FLOW
TIME (RT) RATE
31-4
-------
Introduction to Gas-Liquid Chromatography
i 1 1
SAMPLE 5 iJ.LofStand.inl Pestnulc- Mix-
ture (1 ngo! each Pestici
COM MN Length-O1 X6 mm
Stationary Phase - 10 '„ DC 200
on Anakrom ABS (00 /100 Mesh)
MobilePhn.se- ISOml/minN?
Temperature-2 10°C
5 7 9 11
TIME (IN MINUTES)
Figure 3. GAS CHROMATOGRAM OF PESTICIDE MIXTURE
C Relative Retention Times and Volumes
It is possible to interpret data more easily
by reporting retention data relative to a
particular compound (e.g., aldrin as in
Table 5).
IV QUANTITATIVE ANALYSIS
A Measurement of Peak Area
The quantity of sample component present
is directly proportional to the area under
its peak. (NOTE: This assumption can
only be made if it has been previously
determined that a linear response is ob-
tained in the range under study.) The
following are a few of the ways in which
this area can be measured.
1 Planimeter
2 Calculation of area (see dieldrin peak
in Figure 3)
AREA = peakheight X peak half-width
3 Disc integrator
B Measurement of Peak Height
With the electron capture detector it may
be possible to use peak height for quanti-
tative measurements where the following
conditions are met.
1 A steady baseline is obtained.
2 Retention times can be reproduced
from one injection to the next.
31-5
-------
Introduction to Gas-Liquid Chromatography
Table 5. RETENTION DATA FOR FIGURE 3
Pesticide
Heptachlor
Aldrin
Heptachlor Epoxide
Dieldrin
Retention time (R )
3. 3 minutes
4.2
5.3
7.7
Relative retention
time
0.79
1.00
1.26
1.84
V SUMMARY
The basic components of a gas chromatograph
have been described. Elementary aspects of
quantitative and qualitative analysis are
presented.
BOOKS
Dal Nogare, S., and Juvet, R. S., Jr.
Gas-Liquid Chromatography. New
York: Interscience. 1962.
2 Littlewood, A. B. Gas Chromatograph;
New York: Academic Press. 1962
NEWSLETTERS
1 Aerograph Gas Chromatography News-
letter. Wilkens Instrument and Re-
search, Inc., P.O. Box 313, Walni
Creek, California.
2 F & M Gas Chromatography Newslette
F & M Scientific Corporation. Stan
Road and Route 41, Avondale, Pa.
31-6
-------
POLLUTION PROBLEM OF PESTICIDES
Betty Ann Punghorst*
I NATURE OF PESTICIDES
A Classification
Pesticides include insecticides, herbicides,
rodenticides, miticides, nematicides, and
fungicides. They can be classified accord-
ing to their origin.
1 Mineral origin
These include arsenicals and inorganic
compounds containing sulfur, copper
and fluorine.
2 Botanical origin
These include nicotine, pyrethrum and
red quill.
3 Synthetic origin
a Insecticides
1) Chlorinated hydrocarbons
These compounds all contain
chlorine, hydrogen and carbon
(see Figure 1). A few such as
dieldrin and endrin also contain
oxygen. Otherwise, they vary
widely in their chemical structure
and activity. They affect the
central nervous system; however,
the basic mode of toxic action is
not known for any of them.
2) Organic phosphorus compounds
These compounds are derivatives
of phosphoric acid (see Figure 2)
and are characterized by similar
structure. Their mode of toxic
action is to interfere with
cholinesterase enzyme activity.
b Phenoxyalkanoic acid herbicides
These compounds are similar to
chlorinated hydrocarbons; however,
they are classified separately be-
cause they also contain free carboxyl
groups. Examples of such compounds
are 2, 4D, 2, 4, 5-T and Silvex.
c Miscellaneous compounds (e. g.,
carbamates and dithiocarbamates)
B History of Use and Control
1 In 1000 B. C. , Homer first used sulfur
as a fumigant for pest control.
H H
Cl-C-CI
Cl
DDT
ci
Cl
Cl
Cl
Cl
LINDANE
Figure 1. CHLORINATED HYDROCARBON PESTICIDES
*Chemist, DWS&PC Training Activities, SEC.
WP.PES. 2b. 12. 65
32-1
-------
Pollution Problem of Pesticides
'"'•Ml
P O
CH30
CH30
O
C2H5°
H
C - C - O - C0H
| 2 5
HC - C - O - PH.
25
O
PARATfflON MALATHION
Figure 2. ORGANIC PHOSPHORUS PESTICIDES
2 The insecticidal properties of DDT
were discovered in 1939.
3 Organic phosphorus compounds first
appeared on the market in 1945.
4 The production of synthetic organic
pesticides has skyrocketed from
0 Ibs. in 1942 to 772 million Ibs. in
1963. A ten-fold increase in pesticide
output is predicted in the next 20 years.
5 This large production output of toxic
compounds has necessitated govern-
mental programs of control.
a The original programs were designed
to evaluate mostly short-term effects
(acute toxicity).
1) 1947 - The Federal Insecticide,
Fungicide and Rodenticide Act
required proper labeling of all
formulations containing pesticides.
2) 1954 - The Miller Amendment to
the Federal Food, Drug and Cos-
metic Act of 1938 required that
tolerances be established for
pesticide residues on crops.
b In recent years, concern has arisen
over long-term effects and the
biological magnification of pesticides
in the environment.
C Gains from Use
1 Promotion of health
a Directly through the control of
vector-borne diseases (e. g. , malaria).
II
b Indirectly through increased and
improved agricultural production.
2 Elimination of nuisances.
PESTICIDES AS A WATER POLLUTION
PROBLEM
A History
1 The first awareness that synthetic
pesticides could be a pollution problem
in water occurred in 1950 when extensive
fish kills happened in 14 streams
tributary to the Tennessee River in
Alabama. These kills were caused
by insecticides washed from cotton
fields. <2)
2 In 1954, Tarzwell and Henderson con-
firmed that insecticides could run off
soil with rainwater. They did this by
applying clay granules containing 11%
dieldrin to a measured grassy sod
slope at a rate of 4. 66 Ibs. per acre
and then assaying the dieldrin recovered
in the runoff.(2)
3 In 1959, the PHS established a Pesticide
Pollution Project under the direction of
H. Page Nicholson in Atlanta, Georgia/2'
4 Fish kills occurring in the lower
Mississippi in the winters of 1960,
1961, 1962 and 1963 have been attributed
to synthetic pesticide pollution. Less
than one ppb (water concentration) of
endrin was responsible for the kills.
32-2
-------
Pollution Problem of Pesticides
•B Mode of Entrance into Water
L- 1 Pesticides may be directly applied to
water supplies for one of the following
purposes.
a To control aquatic insects
b To control algal growths
c To poison fish in order to permit
restocking with desired varieties.
__,, 2 Industries may discharge liquid wastes
containing pesticides.
^ 3 Runoff of pesticides from agricultural
lands and forests is dependent on
several factors.
a Solubility of the pesticide in water
b Persistence of the pesticide in the
soil which in turn is dependent on the
pH and temperature of the soil.
c Quantity of pesticide applied to the
soil.
d Formulation of the pesticide
e Method of application of pesticide
f Slope of the land
g Soil characteristics
h Volume and intensity of rainfall
i Soil conservation practices
C Effects
1 Acute toxicity
a An estimated 150 human fatalities
occur annually in the United States
from pesticides. (Note: this is
about the same death rate as that
due to aspirin).
(4)
b Bioassay studies confirm that
many fish are very sensitive to
synthetic pesticides (Table 1).
Table 1. COMPARATIVE TOXICITY OF ORGANIC PHOSPHORUS
AND CHLORINATED HYDROCARBON INSECTICIDES TO
BLUEGILLS IN SOFT WATER
Organic
phosphorus
insecticide
Gluthion
Malathion
Parathion
TEPP
Methyl
Parathion
OMPA
9 6 -hour
TLm
(ppm)
0.0052
0.090
0.095
1.1
1.9
110
Chlorinated
hydrocarbon
insecticide
Endrin
Toxaphene
Dieldrin
DDT
Heptachlor
Lindane
96-hour
TLm
(ppm)
0.0006 f.fb
0.0035
0.0035
0.016
0.019
0.077
Note: Above tests were run under standardized conditions
with soft water as a diluent at 25°C.
32-3
-------
Pollution Problem of Pesticides
2 Chronic toxicity
a In recent years greater concern has
arisen over the possible long term
effects of the accumulation of small
amounts of pesticides in the human
body.
b Another aspect of this problem is
the possibility of synergistic effects
being exerted as these pesticides
accumulate.
3 Taste and odor
Pesticides and the solvents used in
pesticide formulation can be highly
odorous.
Ill SOLUTION TO THE PROBLEM
A More research is needed in order to
answer adequately the following questions:
1 How general is the occurrence of
pesticides in surface and ground
waters? In order to answer this
question, it is necessary that techniques
of analysis be able to measure pesti-
cides in water in the microgram and
nanogramper liter ranges. Pro-
cedures being developed are according
to the following general lines.
a Chlorinated hydrocarbons
1) Recovery of pesticides from
water using the carbon filter or
liquid-liquid extraction.
2) Separation and identification of
extracts.
a) Solubility and chromatographic
separation serves as a clean-
up procedure. Pesticides are
found in the aromatic fraction.
b) Thin layer chromatography
serves both as a clean-up
technique and as a means for
confirmatory identification.
c) Gas chromatography can be
used for identification and
quantitative assay of pesticides.
d) Paper chromatography can
also be used for confirmatory
identification.
3) Infrared identification
Absolute identifications of
pesticides can be made using
micro infrared spectroscopy.
However, it is necessary to
have recovered 1-10 micro-
grams of pesticide when using the
Perkin Elmer Model 421 instru-
ment with 6X beam condenser and
a 1. 5 mm K Br disc.
b Organic phosphorus
1) It is possible to measure organic
phosphorus compounds using
electron capture gas
chromatography.
2)
However, the most popular
techniques developed so far have
been biochemical assays which
measure inhibition of the enzyme
measure inhibition
cholinesterase.^ '
2 What are the less obvious effects of
pesticides on organisms in the aquatic
environment?
3 What factors relate to the presence
or absence of pesticides in water?
4 How well are pesticides removed in
water treatment?
B More Judicious Use of Pesticides
As a result of the special White House
Report (May 14, 1963) and recent Senate
hearings, the Department of Agriculture
has made changes in some of its big spray
projects. Mirex, a compound with low
toxicity to fish, was chosen to replace
heptachlor in the Southeast fire ant
program. Sevin, also a compound with
low toxicity to fish, was chosen to replace
32-4
-------
Pollution Problem of Pesticides
DDT to control the hemlock looper in
Washington State.
IV SUMMARY
The nature of pesticides in general is pre-
sented and their potential as a water pollution
problem is discussed.
REFERENCES
1 Breidenbach, A. W., et al. The Identi-
faction and Measurement of Chlorinated
Hydrocarbon Pesticides in Surface
Waters. PHS Publication # 1241.
September 1964.
2 Nicholson, H. Page. Pesticide Pollution
Studies in the Southeastern States. Pro-
ceedings of the 3rd Seminar on Bio-
logical Problems in Water Pollution.
August 1962.
Pesticides in Soil and Water (an annotated
bibliography). PHS Publication #999-
WP-17. September 1964.
Pickering, Q. H., et al. Toxicity of
Organic Phosphorus Insecticides to
Different Species of Warm water
Fishes. Trans. Amer. Fisheries
Society. 91:175-184. April 1962.
Report on the Use of Pesticides. Prepared
by the President's Science Advisory
Committee Panel on the Use of
Pesticides. May 14, 1963.
Weiss, C. M., and Gakstatter, J. H.
Detection of Pesticides in Water by
Biochemical Assay. J. Water Pol. Control
Fed. 36:240. February 1964.
32-5
-------
BASIC DATA FOR WATER SUPPLY AND WATER POLLUTION CONTROL
Richard S. Green*
I AUTHORITY
A Public Law 410 (78th Congress)
B Presidential Advisory Committee on Water
Resource Policy, 1955 - Recommended
that the existing program of data collec-
tion be accelerated, and carried out on a
more consistent and definitive basis.
C Public Law 660 (84th Congress) - Directs
the Surgeon General, in cooperation with
other Federal, State, and local agencies
to collect and disseminate data on chemical,
physical, and biological water quality and
other information insofar as such data re-
late to water pollution control and pre-
vention.
II NATIONAL INVENTORIES OF WATER
SUPPLY, SEWAGE AND INDUSTRIAL
WASTE FACILITIES
HI FACILITIES CONSTRUCTION AND RE-
LATED ECONOMIC INFORMATION
A Municipal Bond Data - Enables program
personnel to predict probable levels of
future construction.
B Contract Award Data - Shows how munici-
pal funds are being spent for construction
of water supply and sewage facilities.
C Construction "Put-in-Place"Data-Meas-
ures actual progress of construction of
facilities.
D Financing, Operation and Maintenance
Costs, and Related Studies.
E Special Studies of Needs for New Facilities.
IV WATER QUALITY DATA - WATER
POLLUTION SURVEILLANCE SYSTEM
A Municipal Water Facilities
All communities over 100 population -
five year intervals
Communities of 25,000 and over - biennial
B Municipal Sewage Facilities
All communities over 100 population -
five year intervals
C Industrial Waste Facilities
Five year intervals
D Waste Water Disposal at Federal Instal-
lations
A Objectives - to provide:
1 Long term information on changes in
water quality at key points in river
systems as affected by changes in water
use and development.
2 Continuous information on the nature
and extent of pollution affecting water
quality.
3 Data which will guide State, interstate,
and other agencies in their water pol-
lution control programs, and in the se-
lection of sites for legitimate water
uses.
4 Data on water quality useful in the de-
velopment of comprehensive water
resources programs.
E Data Processing and Publication of Inter-
pretive Analyses
*Chief, Basic Data Branch, DWS&PC, Washington, D. C. Reviewed December 1965.
W. 9a. 11.64 33-1
-------
Basic Data for Water Supply and Water Pollution Control
B Sampling Stations Located on:
1 Major waters used for public water
supply, propagation of fish and aquat-
ic life and wildlife, recreational pur-
poses, and agricultural, industrial,
and other legitimate uses.
2 Interstate, coastal, and Great Lakes
waters.
3 Waters on which activities of the Fed-
eral Government have an impact includ-
ing (a) those where a pollution problem
is substantially and adversely affected
by a Federal installation, (b) those
waters needed for national defense,
and (c) those waters involved in a
Federal water resource development.
C Operations
1 Size of System
stations.
- presently 134
2 State and local participation - perform
established analyses and collect sam-
ples for more complex analyses made
in Cincinnati laboratories
3 Compilation, and publication of data -
annual, based on water year from
October 1 to September 30. Statistical
and related studies.
D Laboratory Measurements Include:
1 Long-established examinations - such
as BOD, COD, DO, pH, color, turbi-
dity, temperature, nitrates, chlorides,
alkalinity, hardness, etc.
2 Organic materials - using the carbon
filter technique.
3 Coliform bacteria - using the delayed
incubation, membrane filter technique.
4 Aquatic life - plankton, algae and
bottom organisms.
5 Radioactivity - gross alpha and beta,
strontium-90.
6 Trace elements
E Program Values - Provides:
1 Ultimate measure of effectiveness of
entire water quality control effort.
2 Monitoring of significant changes in
river basin water quality, providing an
"alert", and suggesting possible
control steps.
3 Stimulation of laboratory activities at
local level, encouraging more com-
prehensive coverage and utilization of
water quality parameters needed to
measure the effects of new pollutants.
a The Analytical Reference Service
4 Encouragement and support for intra-
State water quality networks.
5 "Background" levels, required to as-
sess degree of contamination resulting
from the build-up of new pollutants.
V NATIONAL DATA ON POLLUTION-
CAUSED FISH KILLS
33-2
-------
OPERATIONS OF THE WATER POLLUTION SURVEILLANCE SYSTEM
Robert C. Kroner*
I ORGANIZATION OF THE WATER
POLLUTION SURVEILLANCE SYSTEM
II THE FUNCTIONS OF THE SERVICE
LABORATORY ARE:
A The System operation may be considered
to be grouped into 4 separate activities,
namely, the administrative operation,
analytical activities, equipment and instru-
ment development and data utilization.
B Analytical activities are necessarily
devoted to characterization of the samples
and to operations which facilitate the
accuracy of the data. Five different
disciplines are employed for qualifying
and quantifying the samples:
1 Biological examination for algae,
diatoms, other micro- and macro -
plankton and eventually other forms
such as benthos, fish, etc.
2 Microbiological examination for coli-
form and fecal streptococci organisms.
3 Radioactivity assay for alpha and beta
activities in the suspended and dis-
solved fractions, strontium-90 and
eventually other nuclides.
4 Organic materials characterization by
means of the carbon adsorption method.
5 Chemical analysis for 16 conventional
minerals, spectrographic examination
for 16 trace metals and other chemicals.
C Each of the foregoing groups represents
a different discipline and functions as an
independent unit in the System organiza-
tion. The biological, microbiological,
organic and radiological groups examine
samples taken especially for that particular
analysis and obtain all data related to the
sample in the respective laboratory. The
chemical characterization, however, is
not necessarily done by the chemical
analytical group, which leads to the unique-
ness of the Service laboratory.
A When an arrangement is made with a
cooperating agency for participation in the
Water Pollution Surveillance System program,
the agency performs the conventional
analyses as much as possible. Sixteen
mineral determinations are requested, as
follows: Temperature, DO, BOD, pH,
COD, Chlorine Demand (1 hour and 24
hours), ammonia nitrogen, chloride,
alkalinity, total hardness, color, turbidity,
sulfate, phosphate and TDS. Of these 16
measurements, the first 7 named must be
performed by the cooperating laboratory,
the remaining 9 measurements can be,
and in many cases are, performed by the
Service laboratory.
B The fact that the Service Laboratory must
depend upon other personnel of varying
skills and backgrounds to supply data leads
to activities not required by the other
analytical groups in the System organiza-
tion. These activities are:
1 Checking of field data for possible errors
and inconsistencies.
2 Completing analytical work not per-
formed by field stations or repeating
erroneous analyses.
3 Performance of other analyses such as
trace metals by spectrograph.
4 Furnishing standard solutions, reagents,
supplies, etc., to cooperating labora-
tories.
5 Conducting a standard sample program
for cooperating laboratories.
6 Conducting training courses for participa-
ting agencies.
7 Furnishing consultative services, either
personally or by mail.
*In Charge, General Laboratory Services, Water Pollution Surveillance System, SEC.
Reviewed December 1965.
W. LA. 3a. 6. 64
33-3
-------
Water Pollution Surveillance System
HI SPECIFIC ACTIVITIES OF THE SERVICE
LABORATORY
A As the number of cooperating agencies in
the Water Pollution Surveillance System in-
crease, the specific, individual tasks in
the Service Laboratory become more
complex. More field data forms are
handled; characteristics of more stations
require more familiarity; more supplies
are required} personal consultation needs
increase, etc. A brief elucidation of the
more important of these tasks will be
helpful.
1 Checking of field forms. Each coopera-
ting agency reports a weekly sample ,for
conventional mineral analysis. The data
is supplied on the standard forms and
each of the 100 forms is checked weekly
for reporting errors. Erroneous data
is eliminated, questionable data is
checked by analysis whenever possible
and unusual characteristics of the data
are noted.
2 Completing analytical work. Approxi-
mately 25% of the cooperating groups
perform all the 16 required determina-
tions. This leaves a large volume of
analytical work to be performed in the
Service Laboratory and entails con-
siderable bookkeeping on which labora-
tory performs which determinations.
3 Performs other analytical work. A
portion of each weekly sample is
composited until a sufficient volume is
obtained. Supplementary analysis for
sodium, potassium, fluoride, seleni-
um, boron and spectrographic examina-
tion are completed semi-annually.
4 Standard samples. To encourage co-
operating groups to improve their
techniques and to furnish a guide for
reliability of data, standard samples
are supplied to each agency twice a
year. These samples are of two types,
one requiring analysis for conventional
minerals, the other for the demand
type of analyses such as DO, BOD and
COD.
Training courses. In order to increase
the analytical skills of the participating
personnel and to furnish a fuller under-
standing of the System program, train-
ing courses are offered to the station
personnel. The courses at present are
presented semi-annually and consist
of 3 -day sessions devoted to various
phases of the System operations.
B The Service Laboratory and Contract
Services. An important phase of the
Service Laboratory which results from the
assembly of analytical facilities available
in the Water Quality Section is the "con-
tract service" operation. The System
characterizations cover a wide range of
analyses; and because they are geared to
mass production techniques, it is frequently
more efficient for the System laboratories
to perform analytical work for a requesting
organization than it is for the group to
develop its own facilities. This activity,
which is performed on a reimbursable basis
has continued to increase in volume until
at present a large portion of the laboratory
activities are devoted to reimbursable
projects.
It should be noted that these projects in-
volve the organics, planktonic and radio-
logical facilities as well as the chemicals,
but that all contracts are handled through
the Service Laboratory. As a matter of
fact, the term "Service Laboratory" is an
outgrowth of this particular service activity.
33-4
-------
Water Pollution Surveillance System
DEPARTMENT OF HEALTH, EDUCATION, AND WELFARE
PUBLIC HEALTH SERVICE
DIVISION OF WATER SUPPLY AND POLLUTION CONTROL
PROVISIONAL DATA—SUBJECT TO REVISION
FOR WASHINGTON USE ONLY
III)
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33-5
-------
Water Pollution Surveillance System
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33-6
-------
AUTOMATIC INSTRUMENTS FOR WATER QUALITY MEASUREMENTS
D. G. Ballinger*
I NEED FOR CONTINUOUS MONITORING
OF WATER QUALITY
Satisfactory evaluation of the quality of water
depends upon the availability of adequate data.
Such data must be not only as accurate as
economically practical but must also be of
sufficient quantity to support reliable conclu-
sions. In general, the more information
available, the more reliable will be the
interpretations.
In the past, information on the chemical,
physical, and bacteriological quality of most
surface waters has been obtained by periodic
stream surveys or infrequent "spot" analyses.
Although raw water supplies are sampled
daily, the data obtained are restricted to
those tests of importance in water treatment
and do not include other pollutional parameters
or water resources not at present being used
for water supply.
It is apparent that in many situations signifi-
cant changes in water quality may occur often
and abruptly. Seasonal changes in flow, the
occurrence of unpredictable industrial
discharges or spills, and the changes in flow
from impoundments may alter the concentra-
tion of many of the substances of interest to
the water user. In some cases these changes
may occur within a few hours: for example,
the diurnal fluctuations in dissolved oxygen
and the salinity changes in tidal estuaries.
Thus considerable advantage is gained if
continuous monitoring of water quality can be
accomplished. The use of manual sampling
and standard laboratory analyses would be
far too expensive; therefore a wide variety
of automatic instruments have been developed.
II ELECTRICAL INSTRUMENTS
The diagram below illustrates a typical system
for monitoring instruments which utilize
electrical sensors.
The sensor produces an electrical signal
representative of concentration, the signal
ANALYZER
r
1
t
)
J
Figure 1
is converted, and amplified, then passed to
the recorder to develop a permanent record
of concentration levels. For a clear under-
standing of the operational characteristics,
the parts of the system are discussed
separately.
A Sensors
The sensor is the part of the system in
contact with the sample. Sensing
elements may be immersed directly in
the stream, or placed in flow cells through
which the sample is pumped. Both
methods have advantages and limitations.
When the sensor is placed in the stream
the determination is made "in situ. "
Therefore the sample is not affected by
pumping, temperature changes, or time
of travel through the instrument. Such
an installation, however, presents certain
problems. The sensing elements must
be protected from floating debris and
must be mounted so as to remain in a
fixed position in spite of changes in velocity
or direction of current. In bodies of
water which fluctuate in surface level,
such as impoundments, or estuaries, the
relative depth of the sensor may change.
In addition, frequent inspection of the
sensing elements for attached growths or
physical damage is difficult and apt to be
neglected.
The use of the shore-based system, where
the stream sample is pumped through flow
*Supervisory Chemist, Technical Advisory & Investigations Section, Technical Services Branch,
SEC. Reviewed December 1965.
CH. MET. 18a. 9.65
34-1
-------
Automatic Instruments for Water Quality Measurements
cells within the instrument housing, is
free from some of the difficulties mentioned
above. Inspection of the sensors is easy,
cleaning is simplified, and replacement of
sensing elements is readily accomplished.
It should be recognized, however, that the
precautions required for satisfactory
mounting and protection of sensor units
in the stream apply equally as well to the
pump intake. Further, it is essential that
the sample being tested in the flow cell is
truly representative of the stream water.
If dissolved oxygen is included in the
parameters, a submersible pump is
required, to avoid cavitation and prevent
suction removal of dissolved gases. The
intake screen must be carefully designed,
since a fine screen will quickly clog, while
a coarse screen may permit floating
material to enter the system.
Electrical sensors may be conductimetric,
potentiometric, polarographic, or couli-
metric. The sensor may directly measure
a constituent or property of the sample or
it may be used as an indicating mechanism
in automatic titration.
B Analyzer-Amplifier
The function of the analyzer is to convert
the signal from the sensor to a standard
EMF, usually ranging from 0-50 millivolts.
Often bridge circuits are employed. The
analyzer must be rugged to avoid shock
damage and the electronic circuitry must
be stable over a wide range of environ-
mental conditions. Considerable advantage
is gained by the use of standard, readily
available components. Provision should be
made for a standard signal to permit
internal checking of the circuits.
In most cases the signal must be amplified
to provide sufficient voltage to drive a
recorder. Amplification is generally
built into the analyzer circuitry.
C Recorder
Although read-out meters can be used, a
permanent record is desirable. Stripchart
recorders are the most popular even though
they require line voltage. A slow chart
speed is necessary, since observation
periods of several days are common.
Difficulties with pens are often encountered
due to varying conditions of humidity and
temperature. The use of pressure-
sensitive paper may offer distinct advantage.
Circular charts have long been used but
they have certain disadvantages. Unless
the chart is inconveniently large the chart
divisions are very close together making
hourly fluctuations difficult to interpret.
In addition, the circular chart does not
lend itself to mechanical handling of data
for computer processing. Chart drive by
clock mechanism is possible, however,
eliminating the need for power connection.
A recent development in the recording of
data is the use of digital output. Water
quality instruments can be equipped with
analog-to-digital converters which change
the amplitude of the signal to a digital
value. By the use of a punch-tape readout
the data is recorded on paper tape rather
than on a chart. The punched tape may
then be transferred to a computer for data
processing. Therefore, a monitor equipped
with digital tape readout permits the re-
cording of original signal and the per-
formance of statistical computations without
hand transcription.
D Parameters Measured Electrically
The following water quality parameters
can be measured by the use of electrodes:
Temperature
Conductivity
pH
Oxygen
Chloride
Residual chlorine
Oxidation-reduc-
tion potential
A prominent instrument manufacturer is
now conducting research into the develop-
ment of a number of glass electrodes
specifically designed for the measurement
of other ions. It appears that the list above
may be considerably expanded in the future.
34-2
-------
Automatic Instruments for Water Quality Measurements
In most cases precision and accuracy is
not as good as with standard laboratory
tests, but the advantages of continuous
recording outweigh the limitations in
performance.
HI PHOTOMETRIC INSTRUMENTS
Because of the importance of colorimetric
analysis in the water laboratory, considerable
attention has been given to the development of
continuous monitoring instruments employing
this principle. A typical system is shown
in Figure 2.
It should be noted that the sample must be
pumped through the instrument with the
attendant problems described above.
A Measurement of Turbidity and Color
/
In the simplest photometric instrument,
a property of the sample, such as turbi-
dity or color, is measured directly. These
relatively simple parameters, however,
are rather difficult to determine. Turbi-
dity measurements are affected by particle
size and by the true color of the sample.
Conversely, color determinations are
subject to errors caused by turbidity and
by the fact that the wavelength of the color
in the sample may vary widely. Instruments
of this type currently available do not
satisfactorily compensate for these
interferences and the data obtained from
them does not correlate well with standard
methods values.
B Colorimetric Analyzers
A second type of photometric instrument
is designed to reproduce laboratory
colorimetric procedures. That is, re-
agents are added to the sample to produce
a color change proportional to the con-
centration of the material being determined.
Since the sample is flowing continuously
the reagents must be metered accurately
and mixed thoroughly before photometric
measurement. In a properly designed
system, almost any colorimetric procedure
can be duplicated and therefore the potential
range of determinations is much wider than
than in the electrometric instruments.
A modification of the colorimetric instru-
ment is the continuous titrator. In this
system an indicator is added to the flowing
sample and the reagent is added at a vari-
able rate to maintain a constant color.
The amount of reagent required is pro-
portional to the concentration of the re-
actant in the sample, and the current used
by the metering pump acts as a signal for
the analyzer.
ANALYZER
RECORDER
COLORIMETER
SAMPLE
DRAIN
Figure 2
34-3
-------
Automatic Instruments for Water Quality Measurements
In spite of the apparent advantages of the
photometric systems, certain special
problems are inherent. The color and
turbidity of the sample may interfere, the
accumulation of slime in the cells may
seriously reduce the sensitivity, and the
limitations of the filter photometer (wide
band pass) must be considered. Further,
in most colorimetric procedures the
amount of reagent required is proportional
to the concentration range of the sample,
a factor which would limit the applicability
of the instrument.
C Parameters Measured Photometrically
Continuous analytical procedures have
been developed for:
Fluoride
Silica
Phosphate
Phenols
Chemical Oxygen
Demand
Turbidity
Color
Hardness
Residual Chlorine
Alkalinity
IV PERFORMANCE
To illustrate the quality of the data which may
be obtained from an integrated water quality
monitor, the table below shows the results
of a performance test conducted on a proto-
type instrument supplied by one manufacturer.
The "Acceptable" tolerances were selected as
representative of the usual requirements for
continuous data acquisition and may be too
high or too low, depending upon the accuracy
deemed necessary.
V CALIBRATION AND MAINTENANCE
A Calibration
In the parlance of the instrument manu-
facturer, calibration involves two steps:
(1) the setting of the readout to zero value
in the absence of sensor signal, and
(2) setting of the readout to some standard
value, such as 5mv. when the sensor
signal is replaced by a standard signal.
It is readily apparent that a calibration
of this type adjusts the meter and amplifier
to reproduce correctly the signal received
from the sensing elements. It does not,
however, "calibrate" the total instrument
in terms of the concentration of measured
substance in the sample.
For proper calibration, it is essential
that the final readout of the instrument be
adjusted to correspond to a true value for
the measured material. Thus the only
adequate calibration must involve the
measurement of a standard solution, such
as a buffer of known pH or a salt solution
of known conductance. Since instrument
systems may lack linear response to a
wide signal range, the instrument should
be calibrated at several points over the
range of values anticipated. In some
cases current instrument design has not
included adequate means of replacing the
sample with standard solutions for calibra-
tion purposes.
The frequency of recalibration depends
upon both the stability of the analyzer and
the sensor. In general, analyzer-
amplifiers are more stable than the sensing
systems now in use. The PHS National
Test
PH
Temperature
Conductance
Dissolved Oxygen
PERFORMANCE DATA
Acceptable deviation Mean deviation found
+ 0.1 unit
+ 1
0.5mg/l
0.1 unit
0. 2°
0.3 mg/1
% Acceptable
80%
85%
99%
73%
34-4
-------
Automatic Instruments for \\ater Quality Measurements
Water Quality Network, in performance
specifications for monitoring instruments,
has established a two-week period for un-
attended performance. While such instru-
ment stability is desirable, under certain
circumstances, site location and sample
characteristics, may necessitate more
frequent checking and recalibration.
B Maintenance
It is unwise to assume that any stream
monitoring instrument, no matter how
well designed and built, can function
adequately for extended periods of time
without maintenance. Sensing elements
are subject to physical, chemical, and
biological actions, analyzer components
may fail or perform erratically, and
recorders may stop or fail to print. The
frequency of maintenance depends upon a
large number of factors both controllable
and accidental and can only be determined
by long-term testing and actual field
experience. Ease of maintenance can be
designed into the instrument by the use of
replaceable electronic components and
accessibility for cleaning of flow cells
and sample lines.
The availability of trained personnel for
checking and maintaining the instruments
in the field is a key factor in the success-
ful stream monitoring program. Such
personnel need not be either electronic
specialists or experienced analysts but
they should have a working knowledge of
the instrument system and a familiarity
with the chemical principles involved in
the use of standard solutions for
calibration.
VI APPLICATIONS
Stream monitoring systems are now in use
in a variety of locations throughout the
United States. In most cases the instruments
measure a single parameter or condition of
the stream rather than a number of para-
meters. Some of the instruments employ
well established measurement systems, for
example pH and conductance, while others,
such as DO and the colorimetric procedures,
are still largely experimental.
Because of the need for continuous stream
data, the intensive interest on the part of
regulatory agencies, and the development
programs by instrument makers, it is safe
to assume that the use of water quality moni-
toring instruments will increase significantly
in the near future.
34-5
-------
MATHEMATICAL BASIS OF THE BIOCHEMICAL OXYGEN DEMAND (BOD) TEST
D. G. Ballinger*
I FUNDAMENTAL CONCEPTS
Since the early 1900's, the oxidation of
organic waste substances in natural waters
has been under investigation. Certain
fundamentals are now universally accepted.
These are:
A Dissolved oxygen in the water is reduced
during stabilization of the organic material.
B As long as there is dissolved oxygen pre-
sent, the rate of oxidation is independent
of the actual amount of oxygen available.
C The type and numbers of biological forms
present is an important factor.
D Measurement of changes in oxygen content
can be related to quantity and character
of oxidizable organic matter.
II FIRST ORDER RATE CONCEPT
In order to illustrate the mathematical
relationships in the BOD reaction, assume
the following laboratory observations:
A A set of replicate bottles is filled with
river water and sealed so that outside
air is excluded.
B Each day, one of the bottles is analyzed
for DO content, and the results tabulated.
(See Table 1.)
It will be noted that on each successive
day, the DO concentration is less than
the day before. That is, oxygen is being
consumed by biological action in the
water. If the results of the DO test are
plotted against time, a curve, as in
Figure 1 will be produced.
The cumulative oxygen removed from the
sample, (the oxygen demand) when plotted
against time, yields an inverted image
of Figure 1. If the oxygen demand per
day is drawn in, it is apparent that an
increasingly smaller amount of oxygen is
required each successive day, Figure 2.
Note also, a constant percentage of the
oxidizable material present at the begin-
ning of each day is oxidized. Therefore
"K" - the reaction rate coefficient - is
constant from day to day, although it may
Table 1.
Days
DO
Oxygen depletion
Per day Cumulative
0
1
2
3
4
5
6
7
8
9
10
9.2
7.4
5. 8
4. 6
3. 7
2. 9
2. 3
1. 8
1. 5
1. 2
0. 9
0
1.8
1. 6
1. 2
0. 9
0.8
0. 6
0. 5
0. 3
0. 3
0. 3
_
1.8
3.4
4,6
5.5
6.3
6.9
7.4
7. 7
8.0
8. 3
^Supervisory Chemist, Technical Advisory & Investigations Section, DWS&PC, SEC.
CH. O.bod. 49b. 12. 65
35-1
-------
Mathematical Basis of the BOD Test
yv;
/
vary in magnitude from sample to sample,
and with temperature changes.
Similarly, the organic matter present in
the sample is being oxidized, so there is
progressively less oxidizable material
present each successive day. The
relationship between oxygen demand and
^amount of oxidizable material present
can be stated as follows: The oxygen
demand per unit time is proportional to
the amount of unoxidized material present.
Streeter and Phelps^ ' stated the concept
as - the rate of biochemical oxidation of
organic matter is proportional to the con-
centration of unoxidized substance,
measured in terms of oxidizability. The
same authors also stated that there is no
logical reason to expect one rate constant
but it may appear so because of crude
measurement and the effects of many
individual oxidation systems. Such a
rate is termed a First Order Reaction
Rate.
The curve as shown in Figure 2 is the
typical BOD curve. When interpreted in
light of the fundamentals previously dis-
cussed, it is apparent that:
1 With organic materials which are
easily oxidized, the reaction proceeds
rapidly, nearing complete oxidation
in a few days. The curve is therefore
steep, rapidly approaching a maximum.
The slope of the BOD curve is therefore
a function of the rate of oxidation.
2 As oxidation nears completion, the
curve approaches some maximum
BOD value. This value represents
the total oxidizability of the organic
matter in the sample. It is termed
the ultimate demand.
Ill BOD EQUATION
Using the typical BOD reaction curve, it is
possible to develop equations expressing
the various relationships. Labeling the
coordinates as in Figure 3.
t = time in days
L = ultimate demand
L^. = demand remaining at time t
y = demand satisfied at time t
then:
L = y + Lt total demand is equal to
BOD oxidized plus BOD remaining
= fraction of L remaining at time t
I -
L
= fraction of L oxidized at time t
and
t v
1 - — = f- = fraction oxidized
L-i Lj
35-2
-------
Mathematical Basis of the BOD Test
The relationships may be expressed as a
differential equation with respect to "t"
-dL
dt
= KL
(1)
where K is a constant.
Integrating equation (1):
TIME IN DAYS
Lt
Ln_E = -Kt (2)
changing to common logarithms:
L,
t
~~ -kt
(3)
and:
Lt
since — = fraction remaining
L
-kt
then: 10 = fraction remaining
Lt
and since 1 - — = fraction oxidized
then: 1 - 10"kt
L
and: 1 - —^ = 1 - 10"kt
L
= fraction oxidized
- 10"kt (4)
multiplying both sides of equation (4)
by L:
-kt'
L - Lt = L (1 - 10
since: L - L, = y
then: y = L (1 - 10"kt)
;)
(5)
This is the usual form of the BOD
equation. To avoid confusion with
other "k" values, the constant is usually
written as k-^ and the equation becomes
= L (1 - 10~klt)
where:
y = the biochemical oxygen demand
at time t, as measured in the
laboratory.
L = the ultimate demand, when
oxidation is complete.
kj= the logarithmic rate constant.
t = any time interval after oxidation
has begun.
Thus (y) the demand which has been
satisfied at time (t), is dependent upon
the ultimate demand of the organic
material (L), the rate at which
oxidation is taking place (k-j) and the
elapsed time (t).
IV SIGNIFICANCE OF BOD CONSTANTS
kj:The rate is dependent on many factors
such as temperature, nutrients, biological
population, etc. These will be discussed
extensively in another section. Mathemati-
cally, k} is the slope of the BOD curve.
Its effect upon y is shown in Figure 4.
The upper curve represents the BOD of
domestic sewage. The kj rate is 0. 15.
At this rate, 50% of the demand will be
exerted in the first two days, indicating
that the organic matter is readily avail-
able as food for the organisms. In five
days, 83% of the demand has been satisfied
and in 15 days, the oxidation is essentially
complete.
35-3
-------
Mathematical Basis of the BOD Test
The lower curve represents the BOD of
an unpolluted stream. The k-^ rate in this
case is 0. 05. Since the organic matter
present is essentially stable, further
oxidation proceeds slowly. In five days,
only 44% of the demand is satisfied and
at 15 days, there is still 17% of the
material unoxidized. Note that at the
slower rate, it takes 15 days to accomplish
83% oxidation, while at the faster rate,
the same percentage can be accomplished
in 5 days.
EFFECT OF K RATE ON SHORT TERM B.OJD.
Laboratory tests on the BOD of waste
materials are generally 5 day tests.
In the case of these two samples, the
standard 5 day interval would represent
vastly different degrees of oxidation.
Thus it is apparent that 5 day BOD values,
without supplementary knowledge of the
rate of oxidation, are of little practical
value.
L: The ultimate demand L is generally re-
garded as a theoretical limit rather than
an actual one. The conditions which
govern the oxidation reaction are seldom
stable for the extended periods required
to complete oxidation. L values are
useful, however, for predicting the
ultimate demand on the oxygen resources
of the stream. It is the total demand that
is important, not just the demand at a
particular time interval.
Further, since the total demand is
independent of the rate of oxidation,
waste comparisons in terms of L values
are much more significant than comparison
of 5 day BOD values.
Both K and L are approximations useful
for engineering estimates. They are not
"constants". K tends to become smaller
as the more rapidly oxidizing components
of a mixture are exhausted. As K
diminishes the estimate of L increases
for a given y,.
V LOG OF % BOD REMAINING
Using the mathematical relationships
established above, a second BOD equation
can be developed. This equation is useful
when tables of logarithms are available.
since: —— =10 1 = fraction remaining
10~klt X 100 = % remaining
Iog10 (10~klt X 100) = Iog10% remaining
further:
logjQdO'kl* X 100) = 2 - kjt
then:
log of % BOD remaining = 2 - k1t
By the use of this equation, the handling of
exponential values is simplified.
The equations herein developed express the
relationships of the various factors in the
BOD reaction. Since a precise mathematical
arrangement can be shown, the existing
waste problem can be evaluated in terms of
the present oxygen demand, the rate of
stabilization, and the ultimate demand on the
oxygen resources.
REFERENCES
1 Streeter, H. W., and Phelps, E. B.
Public Health Bulletin 46, USPHS. 1925.
2 Phelps, E. B. Stream Sanitation. John
Wiley & Sons, New York. 1944.
35-4
-------
ESTIMATION OF K AND L
F. J. Ludzack*
I Several methods are available that may
give a fair estimate of K and L providing
that the observed data plots in a form that
suggests a first order reaction rate fit.
The fact that stream samples frequently
consist of partially stabilized river water
mixed with varying amounts of more recent
wastewater additions leads to numerous
situations where observed data calculated
as "a" first order reaction does not give a
good fit of observed and calculated data.
The low rate oxidation of the aged material
and high rate oxidation of the new material
simply cannot be explained in terms of one
K and L.
The BOD test is not precise enough to
distinguish the infinite number of individual
rate systems included in deoxygenation of
a mixed wastewater by the mixed organisms
involved. The persistence of the first order
reaction rate concept (incorrectly called a
monomolecular rate) partially is due to the
practical limitations of the test. High
assimilative oxidation rates (Figure 1)
generally have been partially completed
in the sewer, during compositing or sample
storage before BOD analysis, or in the
receiving water. Remaining deoxygenation
takes the form characteristic of cell mass
and storage products, (endogenous oxidation)
which for a few days shows a k^ of about 0.1.
If the observation period was extended beyond
the usual 7 to 10 days for rate estimation a
progressive lowering of k^ would become
apparent as the more oxidizable components
disappeared and relatively inert biological
or other residues became a larger and
larger fraction of the total oxidizable mass.
mathematics as a tool rather than as a deity,
will produce useful estimates for resolution
of the problem.
II Useful methods of deriving K in natural
log or the more commonly used rate term
for deoxygenation, k^ in Iog10 include the
least squares method of Reed and Theriault
(Public Health Bulletin 127, 1927); the
Thomas Slope Method (Sew. Works J. 9,
No. 3, 425, 1937) as modified by Ettinger,
the Moments Method of Moore, Thomas, and
Snow (Sew. and Ind. Wastes 22, No. 10, 1343,
1950) and the Rapid Ratio Method of Sheehy
(JWPCF 32, No. 6, 646, 1960). The first
three of these presume a single rate factor
and do not give a good fit of data when
deoxygenation fails to follow the pattern.
Certain other factors in calculation mechanics
enter the picture even when the curve appar-
ently follows first order characteristics
(Ludzack et al, Sew. and Ind. Wastes 25,
No. 8, 875, 1953). Sheehy's procedure
elucidates the disappearance of high rate
components during early stages and can be
very useful for rapid calculations.
The daily difference method outlined by
Tsivoglou (Oxygen Relations in Streams,
SEC Tech. Report W-58-2, p. 151, 1958)
is an adaptation of Velz' technique and gives
a graphic picture of observed data and pre-
dominant rate changes with time. The method
is rapid, versatile, and gives a great deal
of information in a simple package that is
readily assembled.
A For example with given observed BOD's:
Time interval
Thus the computed k^ and L are a result of
the selected measurement routine, past
history of the sample, and the manner of
processing the result. The validity of the
estimate depends upon engineering judgment
and a realization of the variables and effects
involved. No two individuals will derive
precisely the same values from given data
but common sense approaches using
days
0
1
2
3
4
5
6
7
(mg/1)
0
0.
1.
1.
1.
2.
2.
2.
72
20
55
82
02
20
37
Difference
0
1
2
3
4
5
6
to
to
to
to
to
to
to
1.
2,
3,
4,
5,
6,
7,
0.
0.
0.
0.
0.
0.
0.
72
48
35
27
20
18
17
*Chemist, Chemistry and Physics, Basic and Applied Sciences Branch, DWS&PC, SEC.
CH.O. bod. 55. 12. 65
35-5
-------
Estimation of K and L
SIMULATED HIGH AND LOW RATE
OXIDATION CURVES AND THEIR
INTEGRAL EFFECTS
INCREMENTAL
ADO/AT
ASSIMILATION PHASE
—^=.
^^H^V * ^^^^
^^^** • ^^^^
S ADO vs. Time,
Scale x 1/3
INTEGRAL OR COMPOSITE CURVE
INCREMENTAL
ADO/AT
ENDOGENOUS PHASE
6 8 10
OBSERVATION TIME
Figure 1
12 14
35-6
-------
Estimation of K and L
1 Plot y vs t on coordinate paper. Draw
a curve of best fit through the observed
points, including lags, plateaus, if any,
to get a picture of the deoxygenation
curve. Wild values then can be
eliminated by using the curve values in
subsequent calculations. Lags should
be eliminated by curve fitting and taking
the observed points after the lag
termination.
2 Plot the daily differences, corrected
if necessary, on semilog paper with
time on the linear scale and the daily
difference on the log scale. The
differences are conventionally plotted
as 1/2, 1-1/2, 2-1/2 days, etc.
(Figure 2).
From the plot it is apparent that the
rate changes between the fourth and
fifth day. For calculation of the high
rate system assume that it is no longer
dominant after the fourth day.
3 From 0 and 4 day intercepts of the
line of best fit — ^ or the fraction
23
remaining becomes -~^- = 0. 28. The
fraction oxidized = 1 - - = (1-28)=. 72
L
4 k^ may be estimated from the
Theriault tables where t = 4 and
( 1 - 10"kt) = . 72
where k^ = 0. 14 becomes the nearest
value.
a Alternately k^ may be estimated
from the relation Log of % BOD
remaining = 2 - k^t
where % BOD remaining is 28 and
t = 4
Log 28 = 1. 45
or 1.45 = 2 - ki(4)
kl= °^ =0.14
5 L may be estimated from the Theriault
tables using the relation y^
and known values for yt for 4 days of
1. 82 and (l-10~kt) of . 72
or 1. 82 = L (.72)
Ill From Figure 2 it is clear that the k^ of
0. 14 and L of 2. 5 are subject to the influence
of the lower rate oxidation which is dominant
after the fifth day, but apparently was active
from the initial day. Extending the low rate
line to the initial day provides a means to
estimate the relative contributions of each
rate group.
A The log difference of the corrected
hi-rate group for 0 to 1 day now becomes
. 72 - . 22 = . 50. Subsequent differences
are .27, .15 and .07.
B A new rate estimate corrected for the low
rate system provides a line (Figure 2)
with the 0 intercept at 0. 72 and four day
intercept at 0. 05.
C The % BOD remaining
0 05
becomes - X 100 = 7.
= (lO"kt)
V /
where Log % remaining = 2 - k^t
for four days 0. 85 = 2 - k1(4), or
1. 15
kl =
= 0.29
where yt = L (l-10"kt)
for four days y^. = 1. 82,
(1-10-kt) = (i. 0-0.07) = . 93
L =-:gg- = 1. 96 or 2.0
D The corrected high rate group calculations
now show that:
1 93% of the ultimate rapid rate demand
was exerted in four days.
2 The estimate of L was 2. 5, uncorrected,
and 2. 0, corrected, therefore about 80%
of the observed four day demand was due
to the hi-rate group.
35-7
-------
Estimation of K and L
DAILY DIFFERENCE ADO/AT
V- N» u> :t» cn -^ ootoo o o o o o o o <
St.
vs
\i ^
i \
,,_ | »
1
1
-
-
-
-
\
I x
1
\[
%
-ix
t^ADO
V
V
i
i
i
\
I/AT Ci
ADC
F
\
\
\
)MPO<
ATESJ
I/AT C
1IGHEF
V
SITE
yLOW
* f^. W% i
ORREi
\ RATE
RATE
OUP
;TED
:GROI
-
-i
-
-
-
IP
DAYS
Figure 2
-------
Estimation of K and L
Theriault Table
NUMERICAL VALUES OF THE FUNCTION (1 -
FOR THE RANGE k = 0.040 to k = 0. 250
Period of
incubation
(days)
1__ . __
2-..-....
0 ..
4 -._-«-. -
5 ____-._..-
6 -__.._.._-
7_-__-.-__
8_
9-^---.---
1 9--. _--__-
H ---_-._.
1 ^--_-----
1 A-. ______
1 7--_--__-
1 fl---_-__-
91 _-._--__-
99-.. __--__
9^--_--_-_
9A--------
9R_____-__
Period of
incubation
(days)
2 ______.._
•}--_..____
4 — _
0.04
0.088
.168
.241
.308
.369
.425
.475
.521
.563
.602
.637
.669
.698
.725
.749
.771
.791
.809
.826
.842
.855
.868
.880
.890
.900
0.11
0.224
.397
.532
fi37
0.05
0.109
0.206
0.292
.369
.438
.499
.553
.602
.645
.684
.718
.749
.776
.800
.822
.842
.859
.874
.888
.900
.911
.921
.929
.937
.944
0.12
0.241
.425
.563
fifiQ
0.06
0.129
.241
.339
.425
.499
.563
.620
.669
.712
.749
.781
.809
.834
.855
.874
.890
.905
.917
.928
.937
.945
.952
.958
.964
.968
0.13
0.259
.450
.593
RPQ
Value of k
0.07
0.149
.276
.383
.475
.553
.620
.676
.725
.766
.800
.830
.855
.877
.895
.911
.924
.935
.945
.953
.960
.966
.971
.975
.979
.982
Value of k
0.14
0.276
.475
.620
72Fi
0.08
0.168
.308
.425
.521
.602
.669
.725
.771
.809
.842
.868
.890
.909
.924
.937
.948
.956
.964
.970
.975
.979
.983
.986
.988
.990
0.15
0.292
.499
.645
74Q
. 10 -kt)
0.09
0.187
.339
.463
.563
.645
.712
.766
.809
.845
.874
.898
.917
.932
.946
.955
.964
.970
.976
.981
.984
.987
.990
.991
.993
.994
0.16
0.308
.521
.669
771
0.10
0.206
.369
.499
.602
.684
.749
.800
.842
.874
.900
.921
.937
.950
.960
.968
.975
.980
.984
.987
.990
.992
.994
.995
.996
.997
0.17
0.324
.543
.691
7Q1
35-9
-------
Estimation of K and I,
Period of
(days)
K
6 ...».-. _„_
8 ____..
9 _-__-..-__
1 f)---
n_
1 o_ .._
1 ^--_
U-_ -
1 C _ _ __
1 fi-_-__-__
i 7__,. _
1 R-___«.--.«
1 Q~___ ___
^U
Period of
(days)
i .--._-.-__
3_
4_ _„__
5 __„__--_
6_
7--,.-
8_
9__«. ._„
1 0--------
1 9-.-- --
1 ^__------
1 4__------
0.11
71 fl
7pi
por»
ftfift
PQp
Q91
QOp
QC n
QfJO
Q7 1
. y 1 1
Ql?p
• y i o
QP7
QQA
QQO
• yy^t
0.18
0 339
Q1 7
Q7R
QDA
QQA
QQ7
QQQ
0.12
7AQ
pAQ
pCC
«QH
Q 1 7
QC O
Qfi A
07 o
Q7Q
QQQ
QQ1
QQQ
QQR
0.19
0 354
70.1
QQQ
Q7n
QP.1
QR7
QQ9
QQ7
QQp
QQQ
Theriault Table
Value of k
0.13 0. 14 0.15 0. 16
QAQ Q9J. Q77 QAft
Q79 Q7Q QQA QQQ
.985 .989 .992 .994
QQ9 QQA *.
QQ7 QQQ
Value of k
0.20 0.21 0.22 0.23 0.24
0 369 0 383 0 397 0 411 0 425
602 620 637 653 669
749 766 781 7Q6 809
842 855 868 880 890
900 911 921 929 937
Qfif) Pfifi Q71 97S P79
Q7^ Q7Q Qft? Qflfi QRfl
Q84. QR7 QQfl QQ1 QQ1^
QQO QQP QQ4- QQ.*, QQfi
QQ4. QQ1^ __._ „....-. -_--
QQR 007
QQ7 QQfi -_-» -
QQO QQQ
QQQ QQQ - _- — — .« __..—
0.17
QAC
Q ^c^
QRfi
Q7H
Qpn
Q P7
QQ 1
. yyi
. yyo
.997
"" "* —
0.25
0 438
684
822
900
944
Qfift
QP9
QQO
QQA
QQ7
35-10
-------
Estimation of K and L
Similar calculations for the corrected
low rate group show a k^ of 0. 01 and
L of 7. 6 based on the zero to seven
day extrapolated curve of the low rate
group. The zero and seven day
intercepts show BOD remaining of
. 18
. 22
= . 82 or 82%.
O
z
LU
ee
o
CO
I-
z
UJ
o
cc.
UJ
0.
100
80
60
40
30
20
10
RATES OF B.O.D. SATISFACTION
FOR VALUES OF K, FROM .05 TO.60
14 16 18 20
TIME IN DAYS
35-11
-------
Estimation of K and T,
IV The seven daj observation period gives ,>->plitting in a sludge deposit may produce
enough data to show the basi< characteristics cleavage products showing deoxygenation
of the system. The derived values are rates much greater than 0.01.
relatively crude values based upon specified
conditions and it cannot be assumed that the Derived figures are useful engineering
stream approaches or will maintain those1 estimates for a given system, conditions, and
conditions. For example, the hydrolytic time. They are not absolute or constant values.
35-12
-------
ATOMIC ABSORPTION SPECTROPHOTOMETRY
Nathan C. Malof*
I INTRODUCTION
Atomic absorption is a relatively new method
for determining the presence and amount of
an element in a laboratory sample. In 1953,
Walsh(i) in Australia, recognized its potential
advantage over emission spectroscopy for the
analysis of routine samples. Previously the
method had been used in a restricted way for
analyzing mercury vapor in the atmosphere
and terrestial samples. Walsh devised
apparatus sufficiently simple, versatile and
inexpensive to be applicable to routine analysis
of solutions containing a wide range of elements.
It was not until 1960 that the advantages of
atomic absorption were recognized in the
United States.
II THEORY
If sufficient energy is added to an atom some
of the electrons will move from the normal
or ground state orbital to an orbital of higher
energy or an excited state. Since the higher
energy level is unstable, the electron will
return to its original ground state. The re-
turn to ground state is accompanied by the
release of radiant energy and this energy can
then be measured. This is the principle of
emission spectroscopy. The radiant energy
produces a spectral line of characteristic
wavelength and frequency for a particular
element. An electron will absorb energy
at the same characteristic wavelength at which
it emits energy. This is the basis for atomic
absorption spectroscopy.
In atomic absorption, light from a cathode
made of the element being measured is passed
through a sample which is vaporized by a
flame. The ground state atoms in the flame
absorb the light, diminishing its intensity.
The percent absorption noted by the detector
is then a measure of the concentration of the
element in the sample.
HOLLOW-CATHODE
LAMP
VAPORIZING
ROTATING SYSTEM
CHOPPER v FLAME
| SLIT
MONOCHROMATOR
SAMPLE IN FUEL IN
PHOTO-
DETECTOR
ELECTRONICS
AND
READOUT
Figure 1
*Chemist, Technical Advisory & Investigations Section, DWSPC, SEC.
CH. MET. aa. 1. 12.65 36-1
-------
Atomic Absorption Spectrophotometry
RESONANT
AMPLITUDE A
I <*> ABSORPTION
f
X ABSORPTION
4
u
A
B
ll-XI A
100
D
(l-XI A
E
Figure 2
III INSTRUMENTATION
There are four basic parts to an atomic ab-
sorption spectrophotometer; the light source,
a means of vaporizing the sample, a system
for isolation of resonance line, and the
detector.
A Light Source
For the more volatile elements such as the
alkali metals, mercury and thallium, the
most convenient source is the spectral vapor
lamp.
For line sources of the less-volatile
elements, hollow-cathode discharge tubes
have been found most satisfactory.
B Vaporization of Sample
1 Atomic-absorption methods have been
applied amost exclusively to the analysis
of solutions. For this purpose flames
similar to those used in flame photo-
metry are used.
Several different gas mixtures have
been used such as those shown in Table 1
36-2
-------
Atomic Absorption Spectrophotometry
but for most routine work an air-acety-
lene has been found to be superior to
other mixtures if both convenience and
sensitivity are considered. However,
Al Be, Ti and V, because of the re-
fractory nature of their oxides, prevent
production of sufficient free atoms and
a high temperature flame must be used.
Nitrous oxide produces a hot flame and
presents little danger from explosion.
Table 1
Fuel-Oxidant
Approximate
Temp.,°C
Nitrous Oxide - acetylene 3000
Hydrogen - air 2100
Hydrogen - oxygen 2700 - 2800
Acetylene - oxygen 3100
Acetylene - air 2000 - 2200
Propane - oxygen 2700 - 2800
Illuminating gas - oxygen 2800
Cyanogen - oxygen 4900
There are also two types of burners
used, a) "Total Consumption, " b)
"Premix. "
a The total consumption type is used
for flame work, and is shown in
Figure 3, This burner is often used
in a series of three, and by the use
of mirrors the beam is reflected
three to five times through the flame
to increase the path length.
b The premix burner is the type used
by Walsh and co-workers in their
development work. This burner has
a 10 cm X 1 cm slot and the light
path makes only one pass through the
flame (Figure 4). The advantages
are that the flame is narrower, giv-
ing a larger concentrate of sample
in the flame. Since in the premix
burner only the vapor reaches the
flame, light scattering by droplets
and clogging by salts is avoided.
L
FUEL
OXYGEN
t
SAMPLE
Figure 3
C Line Isolation
The use of a line spectrum of the element
being determined, rather than a continuous
spectrum, makes possible the use of
monochromators of low resolving power
or even filters. When a spectral lamp
is used as a light source, it is only
necessary to isolate the resonance line
from neighboring lines of the light source
or vaporized sample.
D Detector
Photo-electric detectors used in atomic
absorption analysis need be no more sensi-
tive than those used in emission analysis,
since in the atomic absorption method, con-
centration of an element is determined by
measuring the reduction in intensity of the
resonance line emitted from a source of
high intensity.
36-3
-------
Atomic Absorption Spectrophotometry
UJ
o
X
v>
>fle
a
1
g
3
WJ
36-4
-------
Atomic Absorption Spectrophotometry
IV COMPARISON OF ATOMIC ABSORPTION
WITH FLAME METHODS
A Sensitivity
B
Sensitivity mg/1
Flame A. A
Aluminum
Antimony
Arsenic
Barium
Beryllium
Bismuth
Cadmium
Calcium
Cesium
Chromium
Cobalt
Copper
Gallium
Gold
Imdium
Iron
Lead
Lithium
Magnesium
Manganese
Mercury
Molybdenum
Nickel
Palladium
Platinum
Potassium
Rhodium
Rubidium
Selenium
Silver
Sodium
Strontium
Tellurium
Titanium
Thallium
Tin
Vanadium
Zinc
Precision
1 Precision
2
0.3
25
2
0.003
0. 1
0.01
0.2
2
0.002
0. 1
0.01
10
0.001
0.05
0.002
0.01
200
of a single-beam
0.5
0.2
1.0
1.0
0.05
0.2
0.01
0.01
0.05
0.01
0. 15
0.005
1.0
0. 1
0.5
0.05
0. 15
0.005
0.003
0.01
0.5
0.2
0.05
1.0
0.5
0.005
0.3
0.02
1.0
0.02
0.005
0.02
0.5
1.0
0.2
2.0
0.5
0.005
atomic ab-
sorption instrument is primarily a
function of the stability of light output
from the spectral lamp. This in turn is
dependent on the stability of the main
supply and inherent stability of the
lamp. The largest fluctuations are
only + 2 percent for the hollow cathode
tube and sodium spectral vapor lamp.
A double-beam instrument significantly
reduces this error.
2 In common with flame-emission methods,
atomic absorption is subject to "noise"
from the flame and the detector. Changes
in absorption caused by fluctuations in
flame temperature are much less than
those in emission because the strength
of the absorption line varies only as
T 1/2, whereas the intensity of emission
from the flame is much more sensitive
to temperature.
C Accuracy
This is shown by the types of interference
found in flame emission and atomic ab-
sorption spectroscopy. There are three
types:
1 Physical
Collision of atoms and electrons or
atoms and molecules will transfer
energy thus causing an enhancement or
depression of analysis-line emission.
This has a large effect on flame emission
analysis but has only a negligible effect
on atomic absorption.
2 Radiative
Light from elements other than the one
being measured pass the line isolating
device (monochromator or filter). This
occurs in flame emission work, for
example, the interference of calcium
and magnesium in sodium determinations.
This interference is also encountered in
atomic absorption using a D. C. system
but is very small because of the large
signal from the hollow-cathode tube.
Radiative interference is eliminated in
an A. C. system.
3 Chemical
Emission from an element in the flame
is depressed by the formation of
36-5
-------
Atomic Absorption Spectrophotometry
50 100 150 200 250 300 350 400
METAL, ppb
Figure 5
compounds, which are not dissociated
at flame temperatures. This also
affects absorption because the formation
of temperature - stable compounds
in the flame causes proportionate re-
duction in the population of ground-state
and excited atoms.
V REMOVAL OF INTERFERENCES AND
CONCENTRATION OF SAMPLE
A Removal of Interferences
1 The methods for overcoming these inter-
ferences in atomic absorption are similar
to those used in flame emission, namely,
either separation of interfering ions or
suppression of the interference by ad-
dition in excess of a substance that will
prevent formation of compounds between
interfering ions and the element being
determined.
B Concentration of Sample
1 Organic separations can be used to con-
centrate a sample. Interferences are
removed, as seen above, and also the
organic solvent enchanses the absorption.
2 Ion exchange has also been used
successfully for concentrating samples
for atomic absorption.
VI INSTRUMENTS AVAILABLE
A Perkin Elmer
1 Model 303 - double beam, AC - $5, 920. 00
2 Model 290 - single beam, AC - $2, 900. 00
B Beckman attachments for existing
spectrophotometers.
1 Use with model D. U. and D. U. -2 -
single beam, DC - $2, 135. 00
2 Use with model D. B. - single beam,
AC - $2,495.00
C Jarrell-Ash
1 Dual atomic absorption flame spect-
trometer - single beam, AC - $5, 800.00
D E. E.L.
1 Atomic absorption spectrophotometer -
single beam, AC $2, 850.00.
36-6
-------
Atomic Absorption Spectrophotometry
REFERENCES
1 Walsh, A. Spectrochim. Acta. 7, 108. 1955.
2 Kahn, Herbert and Slavin, Walter. Atomic
Absorption Analysis. International
Science and Technology. November 1962.
3 David, D. J. The Application of Atomic
Absorption to Chemical Analysis. The
Analyses. 85:779-791. 1960.
4 Platte, J. A., and Marcy, V.M. A New
Tool for Water Chemicals. Industrial
Engineering. May 1965.
5 Biechler, D. G. Determination of Trace
Copper, Lead, Zinc, Cadmium, Nickel,
and Iron in Industrial Waste Water by
Atomic Absorption Spectrophotometry
After Ion Exchange on Dorvex A-l.
Analytical Chemistry, 37:1054- 1055.
1965.
6 Elwell, W. T., and Gidley, J. A. F. Atomic-
absorption Spectrophotometry. The Mac-
Millan Company. N. Y. 1962.
36-7
-------
AUTOMATION OF CHEMICAL ANALYSIS
Lawrence J. Kamphake*
I NEED FOR AUTOMATED ANALYSIS
A At least 30 samples per week for any
particular analysis.
B The more complex and manipulating time
required for manual method the greater
the advantages to automate.
C Simple manual methods requiring one
titration or instrument reading may be
faster manually unless there are more
than 100 samples per work day.
H DEGREE OF AUTOMATION
A Maximum automation - direct from sampl-
ing source to print-out of results of
multiple analysis.
B Medium automation - individual samples,
instrumental analysis, recording and
calculating data.
C Minimum automation - simplicity and
efficiency in manual analysis.
Ill PRESERVATION OF SAMPLES
A Biological and Chemical Stability
1 Store at 5°C, quick freeze
2 Low pH
3 Chloroform 1%
4 Mercuric chloride 100 mg/1
5 Formalin 500 mg/1
6 Potassium cyanide 50 mg/1
7 Thymol 250 mg/1
IV AUTOMATED INSTRUMENTATION
A Sensory
1 Titrimetric
2 Potentiometric
3 Photometry - flow through cells
a Visible range
b Ultra violet range
c Flame
4 Polarographic - DO
5 Others as coulometric, conducto-
metric, etc.
V MANUAL SYSTEM WITH INSTRUMENTS
A Automatic pipettes and burettes and
reagent dispensers
B Electrical on and off timers
C Glassware, test tubes vs. volumetric
flasks
D Small volumes
E Combining reagents
VI CONTINUOUS FLOW SYSTEMS
A Pumping Systems
1 Peristalic
2 Piston
3 Vibrator
B Reagent Addition
C Removal of Interferences
1 Filtration
2 Dialysis
*Chemist, Engineering Section, Basic and Applied Sciences Branch, DWSPC, SEC. Reviewed
December 1965.
CH. MET. 23. 12.64
37-1
-------
Automation of Chemical Analysis
3 Distillation
4 Chemical complexation
5 Extraction
D System Hydraulics
1 Tubing and glassware
2 Mixing
3 Temperature control
4 Time control
5 Digestion
VI RECORDERS
A Meters
B Chart paper, single pen and multi pens
C Print-out data
VH TECHNICON CORP. - AUTO-ANALYZER
A Equipment and Principle
B Adaption of Standard Methods to
Auto-analyzer
1 Sensitivity
2 Wash-out
3 Chemical reaction and control
C Methods currently in use
1 ABS
2 Ammonia nitrogen
3 Nitrite and nitrate nitrogen
4 Phosphate
D Advantages of Auto-Analyzer
1 Speed - automatic clocks
2 Small volume of sample required
3 Removal of human error
a Timing reactions
b Reading spectrophotometers
c Error in pipetting, recording, etc.
4 Permanent record of data
5 Ease of checking every 10 or 20
samples against standards
6 All samples treated exactly the same
7 Use of chemical methods which
cannot be used manually.
E Disadvantages
1 Acceptable manifold
2 Loss of accuracy in determination of
a low concentration when proceeded
by a high concentration (50 fold)
3 Wash-out difficulty
4 Mechanical failure
5 Pump tubes
F Gradient dilution technique
G Influent and effluent pumped at same rate
-rt
Ct = C1 (1 - e -v- )
C = cone, of influent
Ct = cone, effluent at t time
t = time (min.)
r = pumping rate (ml/min)
v = volume in flask
37-2
-------
Automation of Chemical Analysis
H Simultaneous Analysis of Nitrite and
Nitrate
1 History and chemistry
2 Manifold
3 Variables investigated
4 Accuracy and precision
REFERENCES
1 Muller, R. H. Fully Automatic Titrator
Has Varied Applications. Anal. Chem.
29, 61 A. 1957.
2 Patterson, G. D., Jr. Automatic Opera-
tions in Analytical Chemistry. Anal.
Chem. 29. 605. 1957.
3 Ibid. Anal. Chem. 31. 646. 1959.
4 Ryland, A. L., Pickhardt, W. P. , and
Lewis, C.D. Automation Techniques
in Analytical Chemistry. E.I. duPont
de Nemours & Co. Wilmington, Del.
5 Kamphake, L. J. Simultaneous Analysis
of Nitrite and Nitrate in Water and
Sewage. In Press, R. A.T. Sanitary
Engineering Center. Cincinnati, Ohio.
6 Automatic Chemical Analysis. Annals of
the New York Academy of Science.
Vol. 87. Art 2. 1960.
37-3
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ANALYTICAL REFERENCE SERVICE
Earl F. McFarren*
I INTRODUCTION
The Analytical Reference Service designs and
conducts cooperative studies as a means of
evaluating laboratory methodology in the
field of the environmental sciences and
engineering. Identical samples are dis-
tributed to the member agencies desiring to
participate in any particular study. Inter-
pretation of the data resulting from such
analyses, together with critiques of the
methods employed, provide a basis for such
evaluation. Initially directed toward exam-
ination of water, the studies have been
broadened to include other media.
The Service provides a means of communi-
cation among an increasing number of
agencies whose investment in laboratory
operations is large. For these agencies it
also provides an opportunity for develop-
mental study of newer procedures commen-
surate with the demands of our increasingly
complex environment. The information
resulting from these studies is being utilized
more and more by groups responsible for
establishing standard methods.
II MEMBERSHIP
Membership in the Analytical Reference
Service is voluntary and open to all agencies
having laboratory activities in the field of the
environmental sciences and engineering. It
includes Federal, state, municipal, univer-
sity, industrial and foreign agencies. The
current membership of approximately 205
agencies includes 27 foreign members.
in OBJECTIVES
Through this organization, the cooperative
efforts of the Analytical Reference Service
are directed toward the following objectives:
Statistical evaluation of procedures,
including precision and accuracy.
Comparison of analytical procedures
and results between laboratories having
similar responsibilities.
Exchange of information regarding
method weaknesses.
Improvement of existing methods or
development of newer, more accurate
methods to replace current procedures.
Development and evaluation of entirely
new methodology for the determination
of new pollutional components, such as
pesticides, trace elements, heavy
metals and air contaminants.
IV OPERATION
Studies are selected on the basis of their
importance, of the need to evaluate labora-
tory methods, the availability of satisfactory
procedures, requests from member agencies,
and the need of members to re-evaluate
their own laboratory operations.
Samples are designed and prepared to contain
measured amounts of selected constituents
in the range of concentrations normally
encountered in the environment. Decisions
as to qualitative makeup are made by the
ARS staff, the membership, and consultants.
Notice of each study is forwarded to members
with information regarding the qualitative
makeup and special instructions for sample
treatment. Portions of the sample are
shipped to those members who indicate a
desire to participate in the study. Accom-
panying data forms provide space for numeri-
cal analytical values and a narrative critique
of the procedures used. Laboratories are
urged to comment on the methods, difficulties,
modifications, sources of error, and other
factors.
*Chief, Analytical Reference Service, Training Program, SEC.
CH. ARS. If. 12.65 38-1
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Analytical Reference Service
V STUDIES COMPLETED
Water Minerals Calcium, magnesium hard-
ness, sulfate, chloride, alkalinity, nitrite,
nitrate, sodium and potassium.
Conducted in 1956 - 19 Participants
Conducted in 1958 - 31 Participants
Conducted in 1961 - 69 Participants
Water Metals Lead, copper, cadmium,
aluminum, chromium, iron, manganese and
zinc.
Conducted in 1957 - 16 Participants
Conducted in 1962 - 66 Participants
Water Metals Lead, copper, cadmium,
aluminum, chromium, iron, manganese,
zinc and silver.
Conducted in 1965 - 79 Participants
Water Fluoride Fluoride in the presence and
absence of interferences with and without
distillation, using a specified procedure.
Conducted in 1958 - 27 Participants
Conducted in 1961 - 53 Participants
Water Radioactivity Determination of gross
beta activity.
Conducted in 1959 - 25 Participants
Conducted in 1961 - 41 Participants
Water Radioactivity Determination of Gross
beta and Strontium-90 activity.
Conducted in 1962 - 80 Participants
Water Surfactant Surfactant in various
waters.
Conducted in 1959 - 42 Participants
Conducted in 1963 - 82 Participants
Water Oxygen-Demand Biochemical oxygen
demand and chemical oxygen demand.
Conducted in 1960 - 35 Participants
Conducted in 1964 - 78 Participants
Air Inorganics Chloride, sulfate, fluoride,
and nitrate in aqueous solution and on glass
fiber hi-vol filter mats.
Conducted in 1958 - 20 Participants
Air Lead Lead on filter paper tape. Sample
was designed to approximate pollution from
motor vehicles.
Conducted in 1961 - 42 Participants
Air Participates Microscopic identification
of some common atmospheric particulates.
Conducted in 1964 - 28 Participants
Air Sulfur Dioxide Determination of sulfur
dioxide in air using a specific method.
Conducted in 1962 - 43 Participants
Water Trace-Elements Arsenic, boron,
selenium, and beryllium.
Conducted in 1962 - 29 Participants
Water Trace-Elements Arsenic, boron,
selenium, beryllium and vanadium.
Current study - 92 Participants
Milk DDT - Residue
Conducted in 1962 - 14 Participants
Freshwater Plankton Evaluation of the pre-
cision and accuracy obtainable by using
various methods of plankton counting and
analysis.
Conducted in 1963 - 47 Participants
Pesticides in Milk Dieldrin, DDE, heptachlor
epoxide and lindane.
Conducted in 1965 - 42 Participants
Pesticides in Water Dieldrin, DDE, heptachlor
epoxide and lindane.
Conducted in 1965 - 29 Participants
38-2
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Analytical Reference Service
. VI REPORTS VII PROJECTED STUDIES
The results and comments of each study are Studies in the developmental or proposed
compiled into a comprehensive report which stage include water nutrients and an air
is distributed to all members. It is this report inorganics or sulfur dioxide study.
which provides the means of comparison and
evaluation of the results and techniques.
38-3
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