EPA-430/1-77-003
Self-Monitoring Procedures:
Basic Parameters
for Municipal Effluents
STUDENT REFERENCE MANUAL
U.S. ENVIRONMENTAL PROTECTION AGENCY
OFFICE OF WATER PROGRAM OPERATIONS

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SELF-MONITORING PROCEDURES:
BASIC PARAMETERS FOR MUNICIPAL EFFLUENTS
This course is designed for the treatment plant
operator or technician who is required to monitor
effluent discharges under a National Pollutant
Discharge Elimination System (NPDES) Permit, and
who has had little or no previous experience in
wastewater analysis or flow measurement.
Parameters included in this course are BOQ5, pH,
Fecal Coliform, Residual Chlorine, Suspended Solids,
and Open Channel Flow. At the conclusion of this
training, the student will be familiar with the
standard test procedure for each parameter, will
have performed each analysis, and will be able to
use a parshall flume or weir to measure effluent
flow. He will also know what equipment and sup-
plies are needed in connection with each procedure.
U. S. ENVIRONMENTAL PROTECTION AGENCY
Office of Mater Program Operations
National Training and Operational Technology Center

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DISCLAIMER
Reference to commercial products, trade names, or
manufacturers is for purposes of example and illus-
tration. Such references do not constitute endorse-
ment by the Office of Water Program Operations, U. S.
Environmental Protection Agency.

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CONTENTS
Title or Description
Determination of Five-Day Biochemical Oxygen
Demand (BODg)
Winkler Determination of Dissolved Oxygen-Azide
Modification
Dechlorination of Samples for Biochemical Oxygen
Demand and Seeding of the Dilution Water
Determination of Dissolved Oxygen Using A
Dissolved Oxygen Meter
Determination of Dissolved Oxygen in Wastewater:
Polarographic Probe Method
pH Determination of Wastewater and Wastewater
Treatment Plant Effluents
Collection and Handling of Bacteriological
Samples from a Wastewater Treatment Facility
Fecal Coliform Test by the Multiple Dilution
Tube Method
Fecal Coliform Test by the Membrane Filter
Method
Calculation of the Geometric Mean of Conform
Counts by the Use of Logarithms
Measurement of Flow in an Open Channel by
Parshall Flume
Measurement of Flow in an Open Channel by
Sharp-Crested Weir
Amperometric Determination of Free and
Combined Residual Chlorine 1n Water
Amperometric Determination of Total Residual
Chlorine in Wastewater
Titrimetric Determination of Total Residual
Chlorine in Wastewater Effluents
Determination of Total Suspended
(Non-F1lterable) Solids, mg/l1ter
Settleable Solids, mg/llter (Imhoff Settling Cone)
Reporting of Self-Monitoring Data
164.1.7.77

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A PROTOTYPE FOR DEVELOPMENT OF
ROUTINE OPERATIONAL PROCEDURES
for the
DETERMINATION OF FIVE-DAY BIOCHEMICAL
OXYGEN DEMAND (BODg)
as applied 1n
WASTEWATER TREATMENT FACILITIES
and in the
MONITORING OF EFFLUENT WASTEWATERS
Developed by the
National Training and Operational Technology Center
Municipal Operations and Training Division
Office of Water Program Operations
U.S. Environmental Protection Agency
CH.O.bod.EMP.le.6.77
Page No. 1-1

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EFFLUENT MONITORING PROCEDURE: Determination of Five-day Biochemical
Oxygen Demand (BOD^)
This operational procedure was developed by:
NAME Charles R. Feldmann
ADDRESS EPA, OWPO, NTOTC, Cincinnati, Ohio 45268
POSITION Chemist-Instructor
EDUCATION AND TECHNICAL BACKGROUND
B.S. - Chemistry
M.S. - Chemistry
1-1/2 years Industrial Chemist
4 years additional Graduate School
4 years college Chemistry Instructor
1-1/2 years DHEW - Air Pollution Program, Chemist
10 years DI - EPA, Chemist-Instructor
.Page No. 1-3

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EFFLUENT MONITORING PROCEDURE: Determination of Five-day Biochemical
Oxygen Demand (BOD,-)
1.	Analysis Objectives:
The learner will determine the five-day biochemical oxygen demand of a
sewage sample.
2.	Brief Description of Analysis:
The sample is diluted with a high quality distilled water containing nutrient
salts and a buffer. Two biochemical oxygen demand (BOD) bottles are filled
with the diluted sample. The dissolved oxygen (DO) content of the first
bottle is determined, and expressed as mg of DO/1 iter. The second bottle
is stored in the dark at 20 C for five days. During the five-day period,
microorganisms in the sample break down complex organic matter in the sample,
using up oxygen in the process. At the end of the five-day period, the DO
content of the second BOD bottle is determined, and again expressed as mg
of DO/1 iter. The depletion in oxygen content, divided by the percent of
sample used (expressed as a decimal fraction) is the five-day biochemical
oxygen demand expressed as milligrams of BOD per liter of sample. BODg is
the symbol for the five-day biochemical oxygen demand.
3.	Applicability of this Procedure:
This effluent monitoring procedure describes the determination of five-day
biochemical oxygen demand. It is not applicable when:
a.	The sample contains caustic alkalinity or acidity.
b.	The sample contains residual chlorine compounds. (In this case, consult
the Effluent Monitoring Procedure on the Dechlorination of Samples for
Biochemical Oxygen Demand and Seeding of the Dilution Water. It covers
only the dechlorination and seeding aspects of the biochemical oxygen
demand determination, and then refers the reader back to this Effluent
Monitoring Procedure.)
c.	The sample contains other toxic substances such as those contained in
plating wastes.
d.	The sample is supersaturated with oxygen.
This procedure was excerpted from Standard Methods for the Examination of Water
and Wastewater, 14th ed., pp. 543-550, 1975.
Page No. 1-4

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EFFLUENT MONITORING PROCEDURE: Determination of Five-day Biochemical
Oxygen Demand (BODg)
Equipment and Supply Requirements
A.	Capital Equipment:
1.	Trip balance, 100 g. capacity
2.	Still, or other source of distilled water
3.	Incubator capable of maintaining a temperature of 20 C + 1 C,
and large enough to hold four 300 ml BOD bottles and a 3 liter
jug or bottle
B.	Reusable Supplies:
1.	Brushes (for cleaning glassware)
2.	Brush (for cleaning balance)
3.	Laboratory apron
4.	Safety glasses
5.	One spatula (medium size)
6.	One distilled water plastic squeeze bottle
7.	One pen or pencil
8.	One notebook (for recording data)
9.	Seven plastic weighing boats (2-3 Inches square)
10.	Sponges (for cleaning of laboratory table tops)
11.	One 3 liter jug or bottle with narrow neck
12.	One powder funnel, about 3 inch diameter
13.	One 1 liter volumetric flask
14.	Four 1 liter glass-stoppered bottles
15.	One 1 liter graduated cylinder
16.	One 2 liter graduated cylinder
17.	One siphon (long enough for use with the 2 liter graduated cylinder)
18.	Four 1 ml volumetric pi pets
19.	One 10 ml volumetric plpet
20.	One 20 ml volumetric plpet
21.	One plunger type mixer (for use with the 1 liter graduated cylinder)
22.	Four 300 ml (+ 3 ml) BOD bottles (see pages 16 and 17)
23.	Equipment for doing a Winkler DO determlnation-azide modification,
see EMP on Winkler Determination of Dissolved Oxygen-Azlde Modification.
24.	One dissolved oxygen meter. See the EMP on Determination of Dissolved
Oxygen Using a Dissolved Oxygen Meter (a Weston & Stack, Model 300) or
the EMP on Determination of Dissolved Oxygen 1n Wastewater: Polarographic
Probe Method (a Yellow Springs, Model 54).
25.	One 2 liter beaker (for preparing cleaning solution)
26.	One 12 Inch stirring rod (for preparing cleaning solution)
C.	Consumable Supplies:
1.	Small wad of cotton (to plug the 3 liter jug or bottle)
2.	8.5 g. of potassium dlhydrogen phosphate, KH2P04
3.	21.75 g of dlpotassium hydrogen phosphate, KgHPO^
4.	33.4 g of dlsodlum hydrogen phosphate heptahydrate, Na2HP04'7H20
5.	1.7 g of ammonium chloride, NH^Cl
6.	22.5 g. of magnesium sulfate heptahydrate, MgSOv 7HpO
7.	27.5 g. of anhydrous calcium chloride, CaCt
2	Page Ho.

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EFFLUENT MONITORING PROCEDURE: Determination of Five-day Biochemical
Oxygen Demand (BODg)
C. Consumable Supplies (Continued)
8.	0,25 g. of ferric chloride hexahydrate, FeCl3-6H20
9.	Reagents for doing a Winkler DO determination-azide modification, see
EMP on Winkler Determination of Dissolved Oxygen-Azide Modification
10.	Reagents for use with a dissolved oxygen meter. See EMP on Determination
of Dissolved Oxygen Using a Dissolved Oxygen Meter (a Weston & Stack, Model
300) or the EMP on Determination of Dissolved Oxygen in Wastewater: Polaro-
graphic Probe Method (a Yellow Springs, Model 54).
11.	Concentrated sulfuric acid, H^SO.
12.	Sodium Dichromate, Na5Cr907
13.	Soap	6 c 1
(Items 11, 12, and 13 are for cleaning glassware. The quantities
needed will therefore vary.)
All reagents should be of high quality. Different chemical manufacturers
may have different ways of indicating a high quality reagent. While no
endorsement of one chemical manufacturer over another is intended, the
following are some designations used in four chemical catalogs to indicate
high quality reagents.
Catalog	Designations
Thomas	Reagent, ACS, Chemically
Pure (CP)
Matheson, Coleman	& Bell Reagent, ACS
Curtin Matheson	Primary Standard, ACS, AR
Scientific, Inc.
Fisher	Certified, ACS
Page No. 1-6

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EFFLUENT MONITORING PROCEDURE: Determination of Five-Day Biochemical Oxygen DemandCBODg)
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
A. Equipment Preparation
1.	Cleaning of
glassware
2.	Balance inspection
1. Clean all glassware and
rinse with distilled water.
1. Check all balances for
cleanliness and proper
operation.
la. Consult the manufacturer's manual supplied with
the balance for assistance in correcting any mal-
functions.
V.A.I.1
(P. 15)
B. Reagent Preparation
1.	Distilled water
2.	Phosphate buffer
solution
1.	Distill 3 liters of water
into a small neck jug
(or large bottle).
2.	Plug the jug with a loose
fitting piece of cotton.
3.	Store the jug at 20°C + 1°C
for 48 hours prior to use,
4.	or aerate the water just
prior to use.
1.	Weigh 8.5 g. of potassium
dihydrogen phosphate,
kh2po4.
2.	Heigh 21.75 g. of dipotassium
hydrogen phosphate, K2HPO4.
3.	Weigh 33.4 g. of disodium
hydrogen phosphate hepta-
hydrate, NagHPO^HgO
la. When preparing solutions, unless otherwise spe-
cified, the term water means distilled water,
lb. Unless otherwise specified, solutions should be
stored in glass stoppered bottles.
2a. Air should be able to pass freely into the jug.
3a. This length of time has been determined simply on
the basis of experience.
4a. Do this by shaking the water in a half-filled jug,
4b. or by using a clean supply of compressed air.
(Be cautious about air jets and motors which
may simply contaminate the water with oil).
la. Use plastic weighing boats for weighing the solids,
lb. Use a trip balance for weighing the solids.

°age No. 1-7

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E-^fjjjjffijyNITQKING PROCEDURE: Determination of Five-Day Biochemical Oxygen Demand (BOD^)
Page No. 1-8
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
B. Reagent Preparation
(continued)
3.	Magnesium Sulfate
Solution
4.	Calcium Chloride
Solution
5.	Ferric Chloride
Solution
6.	Dilution Water
4. Weigh 1.7 g. of ammonium
chloride, NH4C1.
,5. Dissolve the four chemicals
together in about 500 ml.
of water.
6. Dilute to 1 liter.
1. Dissolve 22.5 g. of mag-
nesium sulfate heptahydrate,
MgS0.»7H90, in water and
dilute to 1 liter.
1. Dissolve 27.5 g. of anhy-
drous calcium chloride,
CaCl9, in water and dilute
to 1 liter.
1. Dissolve 0.25 g. of ferric
chloride, FeCl3, in water
and dilute to j liter.
1. Siphon 20°C water to the
2000 ml line in a 2 liter
graduated cylinder.
la. If you prime the siphon, "waste" about 50 ml of
liquid before filling the cylinder,
lb. Do not cause splashing which might create air
bubbles. Allow the water to run down the sides
of the cylinder.
1c. Volumes of dilution water larger than 2000 ml may
be prepared if needed.
Id. If larger amounts of dilution water are needed,
Use one ml of each of the four solutions (B.2.,
8.3., b.4., and B.5.) for each liter of distilled
water.
V.B.6.2.
(p. 18)

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EFFLUENT MUNITQkI'G PROCEDURE: Determination of Five-Oay Biochemical Oxygen Demand (BOD,.)
OPERATING PROCEDURES | STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
B. Reagent Preparation
(continued)
2. Add 2.0 ml each of the
buffer, magnesium sulfate,
calcium chloride, and
ferric cnloride solutions.
2a. Use a 2 ml volumetric pi pet for each solution.
2b. Mix gently with a plunger-type mixer after each
solution is added.
2c. The mixture of the four solutions and distilled
water is called dilution water.

C. Procedure
1.	Blanks
2.	Sample Dilution
1.	Fill two BOD bottles with
dilution water by siphon-
ing.
2.	Stopper the bottles.
1. Thoroughly mix the contents
of the sample container,
measure the sample, and
add it to a 1 liter grad-
uated cylinder.
la. Rinse the siphon if it's the same one used above.
lb. Hold the siphon about 1/2 inch from the bottom of
the bottle before opening the siphon.
1c. Open the siphon slowly.
Id. As the liquid level rises in the bottles, keep the
end of the siphon about 1/2 inch above the liquid.
le. Allow a few ml of liquid to overflow the top of
the BOD bottles.
2a. Do not cause formation of an air bubble by insert-
ing the stopper too vigorously.
2b. These two bottles are called blanks.
2c. The DO in one bottle from each pair (two bottles
will be filled for each sample dilution) and in
one blank bottle must be determined within 15 min.
of filling, so proceed immediately to C.2., C.3.,
and C.4. below.
la. Use the appropriate size graduated cylinder to
measure the sample volume.
lb. Pour the sample down the sides of the graduated
cylinder.
1c. During the five-day incubation period, there must
be a depletion of at least 2 mg of DO/1, and at
least 1 mg of DO/1 must remain.
Id. In order to meet these two requirements, it may be
necessary to set up two or three dilutions of each
sample. Use a separate 1 liter graduated cylinder
for each dilution.
V.C.1.1.
(P. 18)
3age No. 1-9

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EFFLUENT MONITORING PROCEDURE: Determination of Five-Day Biochemical Oxygen Demand (BOD^)	Page No. 1-10
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
C. Procedure (Continued)
3. BOD bottle filling
2.	Siphon in dilution water
to the 1000 ml line.
3.	Mix gently with a plunger-
type mixer.
1. For each sample dilution
set up, fill 2 BOD bottles,
by siphoning, from the
liter cylinder.
le. Following, are some suggested sample volumes, al-
though the actual amount of sample to use for each
kind of waste may have to be determined on the
basis of experience.
For effluents of primary treatment plants treating
domestic wastewater:
10.0 ml (1% of the liter volume)
20.0 ml (2% of the liter volume)
40.0 ml (4% of the liter volume)
For effluents of secondary treatment plants treat-
ing domestic wastewaters:
40.0 ml (4% of the liter volume)
60 ml (6% of the liter volume)
80 ml (8% of the liter volume)
For effluents from secondary treatment plants:
200 ml (20% of the liter volume)
300 ml (30% of the liter volume)
400 ml (40% of the liter volume)
If. After experience is gained, it will be necessary
to use only one dilution for each kind of waste,
lg. Measure the sample volumes with an appropriate
size graduated cylinder; e.g., 10 ml size for
10 ml of sample, etc.
2a. Rinse the siphon if it's the same one used above.
2b. Use the same technique as in B.6.1.1a. and lb.
above.
la. Use the same technique as in C.l.la. through le.
above.

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EFFL.UE.Nf MONITORING PROCEDURE: Detsrmination of Five-Day Biochemical Oxygen Demand (BOOc}
OPERATING PROCEDURES
STEP SEQUENCE
INFORMAT1ON/OPE RATIN G GOAL S/S P E CIFICATION S
TRAINING
GUIDE NOTES
C. Procedure (Continued)
4, DO Determination
2. Stopper the BOD bottles.
1.	Fill the flared top of one
of the blank bottles, and
one of the two bottles
filled for each sample
dilution, with water.
2.	Store them at 20°C in the
dark for 5 days.
3.	Determine the DO of the
2nd blank bottle and the
other sample bottle.
4.	Record the results.
5.	After 5 days, determine
the oxygen content of the
stored sample and blank
BOD bottles.
6.	Record the results.
2a. Do not cause formation of an air bubble by in-
serting the stopper too vigorously.
2a. Check the flared tops daily and refill with water
if necessary.
3a. Use the Winkler method-azide modification, or a
dissolved oxygen meter.
3b. These DO values are called initial DO values.
3c. Remember the 15 min. mentioned above.
4a. Page 1-13 is ar. example data sheet.
5a. Use the same method as before.
6a. Page 1-13 is an example data sheet.
V.C.4.2.
(P. 19)
D, Calculations
1. Sample
1. For each pair of sample
bottles, subtract the 00
(in mg/1) after five days
from the initial DO (in
mg/1).
la. Example calculation:
7.0 mg/1 = initial DO
2.0 mg/1 = DO after 5 days
5.0 mg/'i = DO depletion during the 5 days.

3age No- *-11

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EFFLUENT MONITORING PROCEDURE: Determination of Five-Day Biochemical Oxygen Demand (BOD^)
Page No. "1-12
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
D. Calculations
(Continued)
2. Divide the depletion by
the % of sample used (ex-
pressed as a decimal) to
get the B0D,j in mg/1.
2a. Example calculation continued:
The sample was 40 ml of a primary treatment
plant effluent.
The 40 ml were diluted to 1000 ml with
dilution water.
Iqqq x 100 = 4% dilution
4% = 0.04 as a decimal
mg BOD,-/! = 5.0
3 0.04
= 125
2. Blank
3. For the blank BOD bottles
substract the ml of sodium
thiosulfate titrant used
for the stored bottle from
the ml used for the initial
bottle.
3a. The difference is used to calculate a "BOD,-"
for the blank.
3b. Example calculation:
7.5 mg/1 = initial DO
7.3 mq/1 = DO after 5 days
0.2 mg/1 = DO depletion during the 5 days
3c. The blank may be considered as a 100% (1.0 as a
decimal) sample.
3d. mg B0DC/1 for the blank = 0.2 mq/1 =0.2
3	1.0
3e. If it is greater than 0.2 mg/1, the 20°C water
is of low quality.
3f. Possible causes are organic contamination in the
water (check the aeration procedure) or dirty
glassware (especially the BOD bottles and water
storage jug) which has contaminated the water.
3g. This difference is not used as a blank correct-
ion, but merely as a check on the quality of the
20°C water.

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Example Data Sheet
F1ve-Day Biochemical Oxygen Demand (BOD5)
1.	Sample number or other
identification		
2.	ml of sample used		
3.	% dilution (divide line 2 by 10;
assumes dilution to 1,000 ml)		
4.	Decimal egulvalent of %
dilution (move the decimal
point in line 3 two places
left)		
5.	Initial sample DO 1n mg/1		
6.	ml of titrant to titrate
Initial blank		
7.	Date of Initial DO determination		
8.	Time of Initial DO determination			(ADDITIONAL COLUMNS
MAY BE ADDED FOR OTHER
9.	Sample DO after 5 days 1n mg/1			SAMPLE DILUTIONS)
10.	ml of titrant to titrate
5th day blank		
11.	Date of 5th day DO determination		
12.	Time of 5th day DO determination
13.	Line 6 minus line 10*		
14.	Line 5 minus line 9	______
15.	B0D5 in mg/1 (line 14 divided
by T1ne 4)		
~The number here mist not be greater than 0.2.
Page No. 1-13

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TRAINING GUIDE
SECTION	TOPIC
I	Introduction
II	Educational Concepts - Mathematics
III	Educational Concepts - Science
IV	Educational Concepts - Communications
V*	Field & Laboratory Equipment
VI	Field & Laboratory Reagents
VII	Field & Laboratory Analysis
VIII	Safety
IX	Records & Reports
*Training guide materials are presented here under the headings marked *.
Page No. 1-14

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EFFLUENT MONITORING PROCEDURE: Determination <3f Fiye-Day Biochemical
	— Oxygen Demand (B0Dg)
FIELD AND LABORATORY EQUIPMENT
Section v
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
A.1.1
If the glassware 1s especially dirty and cannot be
cleaned with ordinary detergents, chromic acid
cleaning may be required.
1.	Pour 35 ml of distilled water in a 250 ml beaker.
2.	Add about 1/8 teaspoon (simply estimate this
quantity) of sodium dlchromate, Na„Cr?07 to the
water.	c c '
3.	Swirl the beaker until the sodium dlchromate
has dissolved.
4.	Keep repeating steps 2 and 3 until no more
sodium dichromate will dissolve.
5.	Pour the solution into a 2 liter beaker.
6.	Slowly pour 1 liter of concentrated sulfuric
acid, H2S0^, into the 2 liter beaker.
Caution: Use eyeglasses and protective clothing.
7.	Stir the mixture thoroughly.
8.	Store It in a glass-stoppered bottle.
9. The
of about
cleaning solution should be at a temperature
bout 50 C when it is used.
10.	It may therefore be necessary to warm the clean-
ing solution.
11.	When using the warm cleaning solution, fill the
piece of glassware with the solution.
12.	Allow 1t to soak for 2-3 minutes (or longer).
13.	Pour the cleaning solution back Into the storage
bottle.
14.	Rinse the piece of glassware ten times with tap
water.
15.	The cleaning solution may be reused until 1t
turns green.
16.	It should then be discarded.
14th Standard Methods
p. 336, section 2.C.2)
Fafffo 1-15

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EFFLUENT MONITORING PROCEDURE: Determination of Five-Day Biochemical
Oxygen Demand (BODg)
FIELD AND LABORATORY EQUIPMENT
Section V
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
14th Standard Methods does not specify a volume
tolerance for the BOD bottles. A tolerance of
+ 3 ml is suggested by: Methods for Chemical
Analysis of Water & Wastes 1974, U.S. Environmental
Protection Agency. One method of checking the
bottle volume is as follows:
1.	Clean the following items as described in section
A.l, page 7.
a.	One 250 ml graduated cylinder.
b.	One 100.0 ml volumetric pipet.
c.	One 10.0 ml graduated pipet.
d.	One 500 ml Erlenmeyer flask.
2.	Allow the glassware to drain dry.
3.	Turn on the hot and cold water taps at the
laboratory sink.
4.	Adjust the hot and cold water so the temperature
of the water is 20°C. Check it with a
thermometer.
5.	Fill the 500 ml Erlenmeyer flask with the 20°C
water.
6.	Using the 100 ml volumetric pipet, place 300 ml
of the 20°C water in the 250 ml graduated
cylinder. (The meniscus will of course be
above the 0 graduation line.)
7.	Using the 10.0 ml graduated pipet, add 3.0 ml
of the 20°C water to the same cylinder.
8.	Allow the pipet to drain into the sink and shake
It so as to remove water from the tip.
9. Place a mark at the bottom of the meniscus,
is the 303.0 ml graduation mark.
This
10.	Using the same 10.0 ml graduated pipet, remove
10.0 ml of the 20°C water from the 250 ml
graduated cylinder.
11.	Into the sink drain 6,0 ml of water from the
pipet.
12.	Very gently blow the rest of the water 1n the
pipet back Into the 250 ml graduated cylinder.
Page No. 1-16

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EFFLUENT MONITORING PROCEDURE: Determination of Five-Day Biochemical
Oxygen Demand (BODg)
FIELD AND LABORATORY EQUIPMENT
Section V
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
13.	Place a mark at the bottom of the meniscus. This
is the 297.0 ml graduation mark.
14.	Empty the graduated cylinder and allow 1t to
drain dry.
15.	Fill the BOD bottle,whose volume is to be
checked,to overflowing with 20°C water.
16.	Carefully insert the stopper. There must be no
air bubbles in the bottle.
17.	Hold one finger over the stopper and invert the
bottle so as to drain all water from the flared
top.
18.	Hold the bottle upright and carefully remove
the stopper.
19.	Carefully pour the entire contents of the bottle
into the 250 ml graduated cylinder.
20.	If the meniscus is between the 297.0 and 303.0 ml
graduation marks, the BOD bottle may be used. If
the meniscus 1s not, the bottle should not be
used for the BODg test.
Page No. 1-17

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EFFLUENT MONITORING PROCEDURE: Determination of Five-Day Biochemical
Oxygen Demand (BODg)
FIELD AND LABORATORY EQUIPMENT
Section V
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
B.6.2.
If just enough dilution water is prepared for use
at one time, then the procedure in B.6. would be
followed. If you desire to keep a supply of di-
lution water, add 1 ml of the magnesium, calcium,
and ferric solutions for each 1 liter of distilled
water. But do not add the buffer until you are
ready to use the dilution water. The water should
be stored in the dark at 20 C + 1 C.
C.l.l
Standard Methods, 14th ed.
of diluting the sample.
cites two other ways
1. Directly in the 300 ml BOD bottle
Example calculation: A 4% dilution of the
sample is to be made.
300 ml = volume of BOD bottle
.04 = % dilution expressed as a decimal
12.00 = ml of sample to be diluted in the
BOD bottle
Measure the 12 ml of sample, add it directly
to the BOD bottle, and fill the bottle to
overflowing with dilution water.
2. If the dilution will be less than 1%, it should
be done in a volumetric flask.
Example calculation: A 0.5% dilution of the
sample is to be made. Since two BOD bottles
must be filled, make the dilution in a 1 liter
volumetric flask.
1000 ml » volume of flask
.005 * ^dilution expressed as a decimal
5.000 ml 3 ml of sample to be diluted in the
flask
Measure the 5 ml of sample, add it directly
to the flask, and fill to the mark with di-
lution water. Fill the two BOD bottles to
overflowing by siphoning from the flask.
Ho. 1-18

-------
EFFLUENT MONITORING PROCEDURE: Determination of Five-Day Biochemical
Oxygen Demand (BODg)
FIELD AND LABORATORY EQUIPMENT			Section v

TRAINING GUIDE NOTE
REFERENCES/RESOURCES
C.4.2.
There are other ways of keeping the water seal on
the BOD bottle; e.g., the bottles may be turned
upside down in a BOD pan (rectangular pan with
square compartments) half full of tap water or
plastic caps (made for BOD bottles) may be put
over the bottle tops.

MgjC'te. 1-19

-------
A PROTOTYPE FOR DEVELOPMENT OF
ROUTINE OPERATIONAL PROCEDURES
for the
WINKLER DETERMINATION OF DISSOLVED OXYGEN-AZIDE MODIFICATION
as applied in
WASTEWATER TREATMENT FACILITIES
and in the
MONITORING OF EFFLUENT WASTEWATERS
Developed by the
National Training and Operational Technology Center
Municipal Operations and Training Division
Office of Water Program Operations
U.S. Environmental Protection Agency
CH.O.EMP.lC.8.76
Page No. 2-1

-------
EFFLUENT MONITORI FIG PROCEDURE: Winkler Determination of Dissolved Oxygen-
Azide Modification
This process was developed by:
NAME Charles R. Feldmann
ADDRESS EPA, 01JP0, NTOTC, Cincinnati, Ohio 45268
POSITION Chemist-Instructor
EDUCATION AND TECHNICAL BACKGROUND
B.S. - Chemistry
M.S. - Chemistry
1-1/2 years Industrial Chemist
4 years additional Graduate School
4 years college Chemistry Instructor
1-1/2 years DHEW - Air Pollution Program, Chemist
10 years DI - EPA, Chemist-Instructor
Page No. 2-3

-------
EFFLUENT MONITORING PROCEDURE: Winkler Determination of Dissolved Oxygen-
Azide Modification
1. Analysis Objectives:
The operator will be able to perform a Winkler dissolved oxygen
determination, using the azide modification, on a sewage sample.
2.	Brief Description of Analysis:
A solution of manganous sulfate is added to the sample. A solution
containing sodium hydroxide, sodium iodide and sodium azide is next
added. If oxygen is present in the sample, a brown flocculent pre-
cipitate forms. If no oxygen is present, a white precipitate forms.
Sulfuric acid is then added to the sample, and the precipitate dis-
solves. The solution is titrated with sodium thiosulfate using
starch indicator. At the end point of the titration, the color of
the solution changes from pale blue to colorless. The milliliters of
sodium thiosulfate used, is equal to the milligrams of dissolved
oxygen per liter of sample.
3.	Applicability of this Procedure:
This effluent monitoring procedure describes the determination of
dissolved oxygen. The analysis is performed as part of the five-day
biochemical oxygen demand test. Interferences/sample pretreatment
are therefore mentioned in the Effluent Monitoring Procedure,
Determination of Five-Day Biochemical Oxygen Demand (BODg).
This procedure was excerpted from Methods for Chemical Analysis of
Water and Wastes 1974, Methods Development and Quality Assurance
Research Laboratory, National Environmental Research Center,
Cincinnati, Ohio 45268.
Page No. 2-4

-------
EFFLUENT MONITORING PROCEDURE; Winkler Determination of Dissolved Oxygen-
Azide Modification
General Description of Equipment Used in the Process
A.	Capital Equipment
1.	Analytical balance, 200 g. capacity
2.	Trip balance, 500 g. capacity
3.	Oven, temperature control able to + 2°C, large enough to hold a
small evaporating dish
4.	Refrigerator, large enough to hold three 1 liter bottles
5.	Still, or other source of distilled water
B.	Reusable
1.	Hot plate, large enough to hold a 2 liter Erlenmeyer flask
2.	Kemmerer sampler
3.	APHA sampler
4.	Laboratory apron
5.	Safety glasses
6.	Brushes (for cleaning glassware)
7.	Brush (for cleaning balance)
8.	One 1 liter volumetric flask
9.	One 300 ml (+3 ml) BOD bottle (see page 18)
10.	One 1 liter graduated cylinder
11.	One 100 ml graduated cylinder
12.	One 50 ml graduated cylinder
13.	One 10 ml graduated cylinder
14.	Six 1 liter glass stoppered bottles
15.	One rubber stopper (to fit a 1 liter glass stoppered bottle)
16.	One 150 ml glass stoppered bottle
17.	One spatula (medium size)
18.	One spatula (small size)
19.	One 2 liter Erlenmeyer flask
20.	One 500 ml wide mouth Erlenmeyer flask
21.	One 100 ml plpet
22.	One 50 ml pipet
23.	One 20 ml plpet
24.	One plpet bulb
25.	Three 5 ml graduated plpets
26.	One desiccator (large enough to hold a small evaporating dish)
27.	One evaporating dish (large enough to hold about 10 g. of solid)
28.	One 25 ml buret
29.	One ring stand
30.	One buret clamp
31.	One distilled water plastic squeeze bottle
32.	One pen or pencil
33.	One notebook (for recording data)
34.	Eight plastic weighing boats (2-3 inches square)
Page No. 2-5

-------
EFFLUENT MONITORING PROCEDURE: Winkler Determination of Dissolved Oxygen-
Azide Modification
B. Reusable Supplies (Continued)
35.	Sponges (for cleaning of laboratory table tops)
36.	One stirring rod (about 6 inches long)
37.	One powder funnel, about 3 inch diameter
C. Consumable Supplies:
1.	Potassium dichromate, I^C^Oy
2.	Concentrated sulfuric acid, ^04
3.	Soap
(These three reagents are for cleaning glassware. The quantitites
needed will therefore vary.)
4.	480 g. manganous sulfate tetrahydrate, MnSO^HgO
400 g. manganous sulfate dihydrate, MnSO^f^O, or 364 g. manganous
sulfate monohydrate, MnSO^ HgO may also be used.
5.	500 g. sodium hydroxide, NaOH
6.	135 g. sodium iodide, Nal
7.	10 g. sodium azide, NaN.
8.	10 g. soluble starch
9.	15 ml chloroform, CHC13
10.	186.15 g. sodium thiosulfate pentahydrate, Na?S,,0.,-5H,,0
11.	6 g. potassium biiodate, KHUO^
12.	3 g. potassium iodide, KI
13.	10 ml concentrated sulfuric acid, H^SO.
14.	Sodium dichromate, Na2Cr20^
The quantities given in 4 through 11 above will suffice for approximately
450 determinations of dissolved oxygen. Depending on usage, smaller
quantities may be prepared.
All reagents should be of high quality. Different chemical manufacturers
may have different ways of indicating a high quality reagent. While no
endorsement of one chemical manufacturer over another is intended, the
following are some designations used in four chemical catalogs to indicate
high quality reagents.
Catalog	Designations
Thomas	Reagent, ACS, Chemically Pure (CP)
Matheson, Coleman & Bell Reagent, ACS
Curtin Matheson	Primary Standard, ACS, AR
Scientific, Inc.
Fisher	Certified, ACS
Page,; No. 2-6

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EFFLUENT MONITORING PROCEDURE: Winkler Determination of Dissolved Oxygen-Azide Modification
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
A. Equipment Preparation
1.	Cleaning of
glassware
2.	Balance preparation
1. Clean all glassware and
rinse with distilled water.
1. Check all balances for
cleanliness and proper
operation.

V.A.I.1
(p. 17)
B. Reagent Preparation
1.	Manganous sulfate
solution
2.	Alkaline iodide
azfde solution
1. Prepare 1 liter of solution
containing 480 g. of
manganous sulfate tetra-
trydrate, MnSO^-^O
1.	Dissolve 500 g. of sodium
hydroxide, NaflH, in 500 ml
of water.
2.	Cool the solution to room
temperature.
3.	Dissolve 135 g. of sodium
iodide, Nal, in 200 ml of
water.
4.	Dissolve 10 g. of sodium
azide, NaN3» in 40 ml of
water.
5.	Combine the three solutions
and dilute to 1 liter.
la. Unless otherwise specified, solutions should
be stored in glass stoppered bottles,
lb. Unless otherwise specified, the term water means
distilled water.
la. Caution: heat is generated
5a. This solution should be stored in a glass bottle
fitted with a rubber stopper, or in a clean
plastic bottle.

Page No. 2-7

-------
EFFLUENT MONITORING PROCEDURE: Winkler Determination of Dissolved Oxygen-Azide Modification
Page No. 2-8
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
3. Starch solution
1. Gently boil 1 liter of
la. Proceed with the next steps while the water is


water on a hot plate.
heating and boiling.


2. Weigh 10 g. of soluble



starch.



3. Transfer it to a mortar.



4. Add about 3 ml of water.



5. Grind with a pestle so as



to form a thin paste.



6. Pour the paste into the



boiling water.



7. Allow the solution to stand



overnight.



8. Decant the starch solution
8a. Decanting means to pour slowly so that any solid


into a bottle.
material will be left behind.


9. Add 5 ml of chloroform,
9a. Store in a refrigerator.


chci3..

4. Sodium Thiosulfate



stock solution



0.75 N (approximate
1. Boil 1500 ml of water for



3 minutes



2. Cool the water to room



temperature.



3. Weigh 186.15 g. of sodium



thiosulfate pentahydrate,



NagSgOg•5H£0.

-------
EFFLUENT MONITORING PROCEDURE: Winkler Determination of Dissolved Oxygen-Azide Modification
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE MOTES
4.	Continued
5.	Sodium thiosulfate
standard tltrant,
0.0375N
(approximate)
6.	Potassium biiodate
' standard, 0.0375 N
7.	Sulfurte acid,
103! by volume
4.	Oissolve in 500 ml of the
water.
5.	Dilute to 1 liter with more
of the water.
6.	Add 5 ml of chloroform,
CHClr
1.	Dilute 50.0 ml of the sodiui
thiosulfate stock solution
to 1 liter.
2.	Add 5 ml of chloroform,
chci3.
1.	Dry 6 g. of potassium
biiodate, KH(I03)? at
103°C for 2 hours:
2.	Cool in a desiccator.
3.	Prepare 1 liter of a solu-
tion containing 4.837 g. of
the potassiwn biiodate.
4.	Dilute 250.0 ml of this
solution to 1 liter.
1.	Pour 10 ml of concentrated
sulfuric acid, H2SO4, into
90 ml of water
2.	Cool the solution to room
temperature.
6a. Store in a refrigerator.
la. Do not transfer any of the chloroform from the
stock solution.
2a. Store in a refrigerator.
4a. The N of this solution is 0.0375.
la. Caution: pour the acid slowly. Mix after each
addition of 2 ml of the acid.





Page No. 2-9

-------
EFFLUENT MONITORING PROCEDURE: Winkler Determination of Dissolved Oxygen-Azide Modification
Page No. 2-10
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
C. Standardization of
Sodium Thiosulfate
Standard Titrant



1. Potassium biiodate
standard, 0.0375 N
1.	Weigh 1-3 g. of potassium
iodide, KI,
2.	Dissolve in 100-150 ml of
water.
3.	Add 10 ml of 10% by volume
sulfuric acid.
4.	Add 20.0 ml of the 0.0375 N
potassium biiodate.
5.	Place the solution in the
dark for 5 minutes.
6.	Add water so as to bring the
volume to 300ml.
4a. Use a volumetric pipette.

2. Titration
1.	Add the approximately
0.0375 N sodium thiosulfate
titrant to the solution
from a buret until the
color changes from red-brown
to pale yellow.
2.	Add 2 ml of starch solution.
3.	Continue the titration until
the color changes from pale
blue to colorless.
4.	Record the ml of sodium
thiosulfate used.
2a. A medium blue-pale blue color will form.
3a. Ignore any return of blue color.

-------
EFFLUENT MONITORING PROCEDURE: Winkler Determination of Dissolved Oxygen-Azide Modification
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
3. Calculations
1. Divide the ml of sodium
thiosulfate used into 0.75.
la. The result is the normality of the sodium
thiosulfate titrant. It is desirable, but not
necessary, that the normality be exactly 0.0375.
II.C.3.1
(p. 16)
D. Determination of
Dissolved Oxygen
1.	Sample collection
2.	Addition of
reagents
1.	If the sample is to be
collected from a depth
greater than S feet, use a
Kemfterer sampler.
2.	If the sample 1s to be
collected from a depth less
than 5 feet use an APHA
sampler containing a 300 ml
BOO bottle.
3.	If a Kentnerer 1s used,
transfer the sample to a
300 ml BOD bottle. Allow
softe of tiie sample to
overflow.
4.	Carefully Insert the
* stopper of the BOD bottle.
5.	For surface samples, the
sample may be collected
directly in a 300 ml BOD
bottle.
1. Remove the stopper and
pipette 2.0 ml of manganous
sulfate solution Into the
sample.
3a. Caution: during the sample transfer, do not
allow it to splash.
4a. Do not create any air bubbles in the bottle.
5a. Fill the bottle in such a way that no turbulence
is created.
la. Have the tip of the pipette about 1/2 inch below
the surface of the liquid. It is desirable, but
not necessary, that the normality be 0.0375.

Page No. 2-11

-------
EFFLUENT MONITORING PROCEDURE: Winkler Determination of Dissolved Oxygen-Azide Modification
Page No. 2-12
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
2. Continued
2.	Pipette 2.0 ml of alkaline
iodide azide solution into
the sample, over the sink.
3.	Carefully insert the
stopper of the BOD
bottle.
4.	Rinse off the outside of
the BOD bottle.
5.	Holding the hand over the
stopper, invert the BOD
bottle slowly 5 times.
6.	Allow the precipitate to
settle.
7.	Repeat the shaking and
settling steps.
8.	Pipette 2.0 ml of concen-
trated sulfuric acid into
the sample.
9.	Carefully insert the
stopper of the BOD bottle.
10.	Rinse off the outside of
the BOD bottle.
11.	Holding the hand over the
stopper, invert the BOD
bottle slowly five times.
2a. Have the tip of the pipette about 1/2 inch below
the surface of the liquid.
2b. A precipitate forms.
3a. Do not create any air bubbles in the bottle.
4a. The alkaline iodide azide solution is damaging
to the skin.
6a. If it does not settle, wait 2 minutes and proceed.
8a. The pipette need not be below the surface of
the liquid.
9a. Do not create any air bubbles in bottle during
this step.
la. The precipitate will dissolve,
lb. The color of the solution is red-brown if oxygen
is present, but colorless if no oxygen is present.
If the solution is yellow, a small amount of
oxygen is present.

-------
EFFLUENT MONITORING PROCEDURE: Winkler Determination of Dissolved Oxygen-Azide Modification
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
3. Titration
4. Calculations
1.	Transfer the entire contents
of the 300 ml BOD bottle to
a wide mouth 500 ml
Erlenmeyer flask.
2.	Add the sodium thiosulfate
titrant from a buret until
the red-brown color changes
to a pale yellow.
3.	Add '2 ml of starch solution.
4.	Continue the titration until
the color changes from pale
blue to colorless.
5.	Record the ml of sodium
thiosulfate titrant used.
1. Calculate the mg of DO per
liter of sample.
2a. If there was little oxygen in the sample and the
yellow color was therefore present even before
addition of any sodium thiosulfate, the starch
should be added immediately.
3a. A medium blue-pale blue color will form.
4a. Ignore any return of blue color.
la. mg DO/1 iter = ml of sodium thiosulfate
titrant X N of sodium thiosulfate
titrant X 8 x 1000/ml of sample
lb. Since the sample was in a 300 ml BOD bottle,
mg DO/liter « ml of sodium
thiosulfate X N of sodium
thiosulfate X 8 x 1000/300
1c. or,
mg DO/liter = ml of sodium thiosulfate
X N of sodium thiosulfate x 26.7
Page No. 2-13

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EFFLUENT MONITORING PROCEDURE: Winkler Determination of Dissolved Oxygen-Azide Modification
Page no.
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
4. Continued

Id. If the N of the sodium thiosulfate was exactly
0.0375, then
mg DO/fiter * ml of sodium thiosulfate
x 0.0375 x 8 x 1000/300
le. or,
mg DO/liter * ml of sodium thiosulfate x 1

-------
TRAINING GUIDE
SECTION
TOPIC
I
Introduction
II*
Educational Concepts - Mathematics
III
Educational Concepts - Science
IV
Educational Concepts - Communications
V*
Field & Laboratory Equipment
VI
Field & Laboratory Reagents
VII
Field & Laboratory Analysis
VIII
Safety
IX
Records & Reports
~Training guide materials are presented here under the headings marked *.
Page No. 2-15

-------
EFFLUENT MONITORING PROCEDURE: Winkler Determination of Dissolved
Oxygen-Azide Modification
EDUCATIONAL CONCEPTS - MATHEMATICS	Section II
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
C.3.1
The formula used is:
normality of sodium thiosulfate x ml of sodium
thiosulfate = normality of potassium biiodate x ml.
of potassium biiodate.
Three of the four values are known:
ml of sodium thiosulfate is read from the buret.
ml of potassium biiodate = 20.0
normality of potassium biiodate = 0.0375
After rearranging the formula to solve for the
normality of sodium thiosulfate, and inserting the
known values:
Normality of sodium thiosulfate =
20.0 x 0.0375/ml of sodium thiosulfate =
0.75/ml of sodium thiosulfate

Page No. 2-16

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EFFLUENT MONITORING PROCEDURE: Winkler Determination of Dissolved
Oxygen-Azide Modification
FIELD AND LABORATORY EQUIPMENT
Section V
TRAINING GUIDE MOTE
REFERENCES/RESOURCES
If the glassware is especially dirty and cannot be
cleaned with ordinary detergents, chromic acid
cleaning may be required.
1.	Pour 35 ml of distilled water in a 250 ml beaker.
2.	Add about 1/8 teaspoon (simply estimate this
quantity) of sodium dichromate Na2Cr207
to the water.
3.	Swirl the beaker until the sodium dichromate
has dissolved.
4.	Keep repeating steps 2 and 3 until no more
sodium dichromate will dissolve.
5.	Pour the solution into a 2 liter beaker.
6.	Slowly pour 1 liter of concentrated sulfuric
acid, H2SO4, into the 2 liter beaker.
Caution: Use eyeglasses and
protective clothing.
7.	Stir the mixture throughly.
8.	Store it in a glass stoppered bottle.
9.	The cleaning solution should be at a temperature
of about 50°C when it is used.
10.	It may therefore be necessary to warm the
cleaning solution.
11.	When using the warm cleaning solution, fill the
piece of glassware with the solution.
12.	Allow it to soak for 2-3 minutes (or logger).
13.	Pour the cleaning solution back into the storage
bottle.
14.	Rinse the piece of glassware ten times with
tap water.
15.	The cleaning solution may be reused until it
turns green.
16.	It should then be discarded.
14th Standard Methods,
P. 336, section 2.C.2)
Page No. 2-17

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EFFLUENT MONITORING PROCEDURE: Winkler Determination of Dissolved
~	Oxygen-Azide Modification
FIELD AND LABORATORY EQUIPMENT
Section V
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
13th Standard Methods does not specify a volume
tolerance for the BOD bottles. A tolerance of
+ 3 ml is suggested by: Methods for Chemical
Analysis of Water & Wastes 1974, U.S. Environmental
Protection Agency. One method of checking the
bottle volume is as follows:
1.	Clean the following items as described in section
A.1, page 7.
a.	One 250 ml graduated cylinder.
b.	One 100.0 ml volumetric pipet.
c.	One 10.0 ml graduated pipet.
d.	One 500 ml Erlenmeyer flask.
2.	Aliov/ the glassware to drain dry.
3.	Turn on the hot and cold water taps at the
laboratory sink.
4.	Adjust the hot and cold water so the temperature
of the v/ater is 20°C. Check it with a
thermometer.
5.	Fill the 500 ml Erlenmeyer flask with the 20°C
water.
6.	Using the 100 ml volumetric pipet, place 300 ml
of the 20°C water in the 250 ml graduated
cylinder. (The meniscus will of course be
above the 0 graduation line.)
7.	Using the 10.0 ml graduated pipet, add 3.0 ml
of the 20°C water to the same cylinder.
8.	Allow the pipet to drain into the sink and shake
it so as to remove water from the tip.
9.	Place a mark at the bottom of the meniscus. This
is the 303.0 ml graduation mark.
10.	Using the same 10.0 ml graduated pi pet» remove
10.0 ml of the 20°C water from the 250 ml
graduated cylinder.
11.	Into the sink drain 6.0 ml of water from the
pipet.
12.	Very gently blow the rest of the water in the
pipet back Into the 250 ml graduated cylinder.
Page No. 2-18

-------
cFFLUETiT MONITORING PROCEDURE: Winkler Determination of Dissolved
Oxygen-Azide Modification
FIELD AND LABORATORY EQUIPMENT	Section V

TRAINING GUIDE NOTE
REFERENCES/RESOURCES

13.	Place a mark at the bottom of the meniscus. This
is the 297.0 ml graduation mark.
14.	Empty the graduated cylinder and allow 1t to
drain dry.
15.	Fill the BOD bottle,whose volume is to be
checked,to overflowing with 20°C water.
16.	Carefully insert the stopper. There must be no
air bubbles in the bottle.
17.	Hold one finger over the stopper and invert the
bottle so as to drain all water from the flared
top.
18.	Hold the bottle upright and carefully remove
the stopper.
19.	Carefully pour the entire contents of the bottle
into the 250 ml graduated cylinder.
20.	If the meniscus is between the 297.0 and 303.0 ml
graduation marks, the BOD bottle may be used. If
the meniscus is not, the bottle should not be
used for the BODg test.

Page No. 2-19

-------
A PROTOTYPE FOR DEVELOPMENT OF
ROUTINE OPERATIONAL PROCEDURES
for the
DECHLORINATION OF SAMPLES FOR BIOCHEMICAL
OXYGEN DEMAND AND SEEDING OF THE DILUTION WATER
as applied in
WASTEWATER TREATMENT FACILITIES
and in the
MONITORING OF EFFLUENT WASTEWATERS
Developed by the
National Training and Operational Technology Center
Municipal Operations and Training Division
Office of Water Program Operations
U.S. Environmental Protection Agency
CH.O.bod.EMP.2.6.77
Page No. 3-1

-------
EFFLUENT MONITORING PROCEDURE: Dechlorination of Samples for Biochemical
Oxygen Demand and Seeding of the Dilution Water
This operational procedure was developed by:
NAME Charles R. Feldmann
ADDRESS EPA, OWPO, NTOTC, Cincinnati, Ohio 45268
POSITION Chemist-Instructor
EDUCATION AND TECHNICAL BACKGROUND
B.S. - Chemistry
M.S. - Chemistry
1-1/2 years Industrial Chemist
4 years additional Graduate School
4 years college Chemistry Instructor
1-1/2 years DHEW - Air Pollution Program, Chemist
10 years DI - EPA, Chemist-Instructor
Page No. 3-3

-------
EFFLUENT MONITORING PROCEDURE: Dechlorination of Samples for Biochemical
Oxygen Demand and Seeding of the Dilution Water
1.	Analysis Objectives:
The learner will dechlorinate a sample of wastewater treatment plant
effluent, and seed a supply of dilution water for use in the biochemical
oxygen demand test.
2.	Brief Description of Analysis:
Chlorine is added to wastewater treatment plant effluents in order to
destroy undesirable bacteria. Part of the chlorine is used up by chemical
reaction with pollutants in the effluent. The resulting chlorine compounds,
and the chlorine which has destroyed the bacteria, are an interference in
the biochemical oxygen demand test. If the chlorine is not chemically
"neutralized," the biochemical oxygen demand results will be meaningless.
The chemical "neutralization" is accomplished by adding a calculated amount
of sodium sulfite solution to the sample.
Because bacteria in the sample have been destroyed by the chlorination step,
they must be replaced in order to carry out the biochemical oxygen demand
test. This is accomplished by introducing bacteria from domestic sewage
into the water used to dilute the dechlorinated sample.
3.	Applicability of this Procedure:
a.	Theoretically, this procedure can be used to "neutralize" any concentra-
tion of chlorine. In practice, the concentration which would have to be
"neutralized" would probably not exceed 2 or 3 mg/liter.
b.	Any needed preservation techniques would be those required for the
biochemical oxygen demand test itself. Any loss of chlorine, such as
through agitation, or exposure to sunlight, would actually be beneficial,
since there would be less to "neutralize."
c.	If the biochemical oxygen demand sample is taken prior to chlorination,
the "neutralization" and seeding steps described in this procedure are
unnecessary. In this case, the effluent monitoring procedure, Deter-
mination of Five-Day Biochemical Oxygen Demand (BODg), should be used.
Source of Procedure: Standard Methods, 14th ed., par. 4.C.2) page 546
Page No. 3-4

-------
EFFLUENT MONITORING PROCEDURE: Dechlorination of Samples for Biochemical
Oxygen Demand and Seeding of the Dilution Water
Dechlorinate 100 ml of sample
Calculate amount of sodium sulfite
to dechlorinate entire sample
Dechlorinate entire sample
Check completeness of dechlorination
Prepare seed controls
Seed the dilution water
Do Initial and final dissolved oxygen
determinations on the seed controls and
the dechlorinated sample
Final calculations
Page No. 3-5

-------
EFFLUENT MONITORING PROCEDURE: Dechlorination of Samples for Biochemical
Oxygen Demand and Seeding of the Dilution Water
General Description of Equipment Used in the Process
A.	Capital Equipment:
1.	Trip balance, 100 g capacity
2.	Analytical balance
3.	Still, or other source of distilled water
4.	One incubator, 20 C + 1 C (large enough for 6 BOD bottles and a 2 liter
Erlenmeyer flask
B.	Reusable Supplies:
1.	Brushes (for cleaning glassware)
2.	Brush (for cleaning balance)
3.	Laboratory apron
4.	Safety glasses
5.	One distilled water plastic squeeze bottle
6.	One pen or pencil
7.	One notebook (for recording data)
8.	Sponge (for cleaning laboratory table top)
9.	Three 1 liter graduated cylinders
10.	One 500 ml graduated cylinder
11.	One 250 ml graduated cylinder
12.	One 100 ml graduated cylinder
13.	One 50 ml graduated cylinder
14.	One 10 ml graduated cylinder
15.	One 250 ml beaker
16.	One hot plate
17.	One magnetic stirrer (optional)
18.	One magnetic stirring bar (optional; about 1 inch long)
19.	One small spatula (for use when weighing solids)
20.	One 25 ml buret
21.	One small funnel (to fit in the top of the buret)
22.	One clamp (to support the buret)
23.	One ring stand (for use with the buret and clamp)
24.	One mortar and pestle (about 100 ml capacity)
25.	One eyedropper
26.	One 1 ft. long stirring rod
27.	One 10 ml graduated pi pet
28.	One 100 ml volumetric pi pet
29.	One 50 ml volumetric pi pet
30.	Two 1 liter glass-stoppered bottles
31.	Two 100 ml glass-stoppered bottles
32.	Two 2 liter Erlenmeyer flasks
33.	One 500 ml Erlenmeyer flask
34.	One 250 ml Erlenmeyer flask
35.	Four plastic weighing boats (2 inches on an edge)
36.	One Erlenmever flask or large bottle containing about 5 liters of dilution
water at 20 C + 1 C. (See the effluent monitoring procedure, Determination
of Five-Day Biochemical Oxygen Demand, BOD,-, sections B.l. through B.6.
for its preparation.
Page No. 3-6

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EFFLUENT MONITORING PROCEDURE: Dechlorination of Samples for Biochemical
Oxygen Demand and Seeding of the Dilution Water
B.	Reusable Supplies (Continued)
37.	One siphon (long enough to reach to the bottom of the above container)
38.	One siphon (long enough to reach to the bottom of a 1 liter graduated
cylinder, item 37 may be used if it is thoroughly rinsed)
39.	One plunger type mixer (for use with the 1 liter graduated cylinders)
40.	Twelve 300 ml (+3 ml) BOD bottles
41.	One pi pet bulb
42.	One 1 liter volumetric flask
43.	Asbestos gloves or crucible tongs
44.	Equipment for determination of dissolved oxygen by the Winkler Method-
azide modification, or by the use of a dissolved oxygen meter. See the
appropriate effluent monitoring procedure.
C.	Consumable Supplies:
1.	Concentrated sulfuric acid, H2S0., 5 ml
2.	Potassium iodide, KI, 10 g
3.	Anhydrous sodium sulfite, Na^SO-, 2 g
4.	Soluble starch, 5 g
5.	Salicylic acid, CjHgOg, 1.25 g
6.	Reagents for determination of dissolved oxygen by the Winkler Method,
azide modification, or by the use of a dissolved oxygen meter.
All reagents should be of high quality. Different chemical manufacturers
may have different ways of indicating a high quality reagent. While no
endorsement of one chemical manufacturer over another is intended, the
following are some designations used in four chemical catalogs to indi-
cate high quality reagents.
Catalog	Designations
Thomas	Reagent, ACS, Chemically Pure (CP)
Matheson, Coleman & Bell	Reagent, ACS
Curtin Matheson	Primary Standard, ACS, AR
Scientific, Inc.
Fisher	Certified, ACS
Page No. 3-7

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EFTLUiENT_MOr|QiTOiRJ_NG__PiROCiEiD^RF^: Dechlorination of Samples for Biochemical Oxygen Demand and
Seeding of the Dilution Water
Page No. 3-8
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
A. Equipment Preparation
1.	Cleaning of glass-
ware
2.	Balance inspection
1. Clean all glassware and
rinse with distilled water.
1. Check all balances for
cleanliness and proper
operation.
la. Throughout the remainder of this procedure, un-
less otherwise stated, the word water means
distilled water.
la. Consult the manual supplied with the balance if
you are unable to correct any malfunction of the
balance.
V.A.1.1
B. Reagent Preparation
1. Seed material
1.	Collect 1 liter of domestic
sewage influent in a 2
liter Erlenmeyer flask.
2.	Place the flask in an in-
cubator for 24-36 hours.
la. Simply estimate this volume.
2a. At 20°C + 1°C.
2b. There should be no stopper in, or cover on, the
flask.
2c During normal working hours, swirl the flask for
about 1 minute every 2 hours. This will ensure
that the sewage is thoroughly mixed with oxygen
from the air.
2d The solids in the sewage must be completely settled
when the sewage is used in this procedure. There-
fore, the last swirling should be done at least
2 hours before use.
2e. These steps must be done 24-36 hours before the
sample is to be dechlorinated and the BOD set up,
in order to prevent any delay in the procedure.
2f. The supernatant liquid above the solid material
is called seed.
2g.This seed material will be used in sections F.l.
and F.3.

-------
EFFLUENT MONITORING PROCEDURE: Dechlorination of Samples for Biochemical Oxygen Demand and
""~' Seeding of the Dilution Water
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
B. Reagent Preparation



(Continued)



2. Sulfuric acid
1. Measure 50 ml of distilled
la. Use a 50 ml graduated cylinder.

solution, HUSO.,
water.


1+50 24




2. Pour it into a 250 ml



Erlenmeyer flask.



3. Measure 1 ml of concen-
3a. Use a 10 ml graduated cylinder.


trated sulfuric acid,
3b. It will be more convenient, and safer, to pour


HoSO..
2 or 3 ml of the acid into a small beaker, and


C 4
pour it from the beaker into the cylinder. The



excess acid may be discarded.


4. Pour it into the Erlen-



meyer flask.



5. Swirl the flask to mix



the contents.



6. Store the solution in a



100 ml glass stoppered



bottle.


3. Potassium iodide
1. Weigh 10 g of potassium
la. Use a trip balance.

solution, KI, 10%
iodide, KI.
lb. Use a plastic weighing boat.


2. Transfer it to a graduated



250 ml Erlenmeyer flask.



3. Add water to the flask to



the 100 ml mark.



4. Swirl the flask to dissolve



the solid.


Page No. 3-9

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EFFLUENT MONITORING PROCEDURE: Dechlorination of Samples for Biochemical Oxygen Demand and
"""Seeding of the Dilution Water
Page No. 3-10
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
B. Reagent Preparation
(Continued)
4.	Sodium sulfite
solution, Na„SOv
0.025 N c J
5.	Starch indicator
5. Store the solution in a
100 ml glass stoppered
bottle.
1.	Weigh 1.575 g of anhydrous
sodium sulfite, Na2S03.
2.	Transfer it to a 1 liter
volumetric flask.
3.	Fill the flask about half
full with water.
4.	Swirl the flask to dis-
solve the solid.
5.	Add water to the 1 liter
mark.
6.	Thoroughly mix the con-
tents of the flask.
7.	Store the solution in a
1	liter glass stoppered
bottle.
1.	Weigh 5 g of soluble
starch.
2.	Transfer it to a mortar.
3.	Measure 1 liter of water.
4.	Pour the water into a
2	liter Erlenmeyer flask.
la. Use an analytical balance,
lb. Use a plastic weighing boat.
7a. The concentration of this solution is 0.025 N.
7b. It is not stable, and must be prepared fresh
on each day it is used.
la. Use a trip balance.
lb. Use a plastic weighing boat.
3a. Use a 1 liter graduated cylinder.

-------
EFFLUENT MONITORING PROCEDURE: Dechlorination of Samples for Biochemical Oxygen Demand and
"""""Seeding of the Dilution Water
OPERATING PRGCEDURES
STEP SEQUENCE
INFORMAT ION/0PERAT ING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
B. Reagent Preparation
(continued)
5.
Bring the water to a boil.
5a.
5b.
Use a hot plate.
While the water is coming to a boil, do steps
6 and 7.


6.
While the water is coming
to a boil, add 1 ml of
water to the starch in the
mortar.
6a.
Use a 10 ml graduated cylinder.


7.
Grind the starch and water
together.
7a.
7b.
7c.
Use a pestle.
The objective is to form a thin paste.
A few additional drops of water may have to
be added.


8.
Pour the thin paste slowly
into the boiling water.




9.
Invert a 250 ml beaker and
place it on top of the
Erlenmeyer flask.




10.
Turn the hot plate off.




11.
Remove the flask from the
hot plate.
11a.
lib.
Caution: The flask is hot.
Use asbestos gloves or crucible tongs to move
the flask.


12.
Allow the starch solution
to stand overnight.




13.
Carefully decant the super-
natant liquid into a 1
liter glass-stoppered
bottle.
13a.
Recall that decant means to carefully pour out
the liquid and leave any solid material behind.

Page No. 3-11

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EFFLUE^^fWNIKJRUIG^PRQCEDUR^: Dechlorination of Samples for Biochemical Oxygen Demand and
Seeding of the Dilution Water
Page No. 3-12
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
B. Reagent Preparation
(continued)
6. Dilution water
14.	Weigh 1.25 g of salicylic
acid.
15.	Add it to the starch solu-
tion in the bottle.
16.	Swirl the bottle to dis-
solve the solid.
1. Prepare the needed quan-
tity of dilution water.
14a. Use an analytical balance (or trip balance if it
weighs to the second decimal place).
14b. Use a plastic weighing boat.
la. See the effluent monitoring procedure on the
Determination of Five-Day Biochemical Oxygen
Demand, sections B.l. through B.6., for the pre-
paration of the dilution water. Throughout the
remainder of this procedure, the EMP on BOD,- will
be used to mean the effluent monitoring procedure
on the Determination of Five-Day Biochemical
Oxygen Demand,
lb. A maximum of 300 ml is needed for each BOD bottle,
whether it contains a sample, seed material, or
a dilution water blank,
lc. Multiply the number of bottles to be set up by
300 to get the total volume of dilution water
needed. Prepare an extra 500 ml for "safety".
Six bottles are needed for the seed material, and
2 are needed for the dilution water blank. See
the EMP on BOD5, sections C.l. and C.2., for the
number of sample bottles needed.
Id. Recall from the EMP on BOD5 that the dilution
water should be at 20° + 1°C when used. When not
being used, it is stored at this temperature.

-------
EFFLUElirT^WNIT^R^iG^^PROCEDL^R^: Dechlorination of Samples for Biochemical Oxygen Demand and
Seeding of the Dilution Water
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
C. Determination of
amount of Sodium Sul-
fite needed for de-
chlorination
1.	Pi pet 100 ml of well mixed
sample into a 500 ml Erlen-
meyer flask.
2.	Measure 10 ml of the sul-
furic acid solution.
3.	Add it to the Erlenmeyer
flask.
4.	Swirl the flask to mix
the contents.
5.	Measure 10 ml of the potas-
sium iodide solution.
6.	Add it to the Erlenmeyer
flask.
7.	Swirl the flask to mix
the contents.
8.	Fill a 25 ml buret to the
0.0 line with the 0.025 N
sodium sulfite.
9.	While swirling the flask
vigorously, add the sodium
sulfite from the buret.
la. Use a volumetric pipet. If solids clog the tip
of the pipet, use a graduated cylinder.
lb. See Sections C.l and C.2. of the EMP on BOD5 to
determine the amount of sample needed. Since
100 ml of sample are needed to determine the
quantity of sodium sulfite required for dechlori-
nation, the sample volume collected should be
about 100 ml, plus the amount of sample needed for
the BOD5, plus about 200 ml for "safety." Simply
estimate the total volume when collecting the
sample.
2a. Use a 10 ml graduated cylinder.
5a. Use a 10 ml graduated cylinder.
7a. The color of the solution is red brown.
9a. Add the sodium sulfite at a fast, drop-wise rate.
9b. A magnetic stirrer may be used.

Page No. 3-13

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EFFLUENT MONITORING PROCEDURE: Dechlorination of Samples for Biochemical Oxygen Demand and
Seeding of the Dilution Water
Page No. 3-14
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
C. Determination of
Amount of Sodium Sul-
fite needed for de-
chlorination (con-
tinued)
10.	When the red-brown color
changes to a pale yellow
color, stop the addition
of sodium sulfite.
11.	Measure 2 ml of the starch
indicator.
12.	Add it to the flask.
13.	Swirl the flask to mix the
contents.
14.	While swirling the flask
(or using the magnetic
stirrer), begin again to
add the sodium sulfite
solution from the buret.
15.	When the solution turns
from pale blue to color-
less, inmediately stop the
addition of sodium sulfite.
16.	Record the ml of sodium
sulfite used to one place
to the right of the
decimal point.
11a. Use a 10 ml graduated cylinder.
lib. Although the cited reference does not specify
what volume of starch indicator is to be used,
two ml is a commonly used quantity.
13a. The solution will be medium or pale blue in color.
14a. At a rate of about 1 drop per second.
14b. The solution will become lighter blue in color.
14c. Read step 15 before carrying out step 14.
16a. Note that the 100 ml of sample you have just
titrated should now be discarded. The actual
BOD test will be done on another, and larger
portion of sample.
1

-------
EFFLUENT MONITORING PROCEDURE: Dechlorination of Samples for Biochemical Oxygen Demand and
™"""""""""" Seeding of the Dilution Water
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
D. Calculations
1.	Calculate the amount of
the 0.025 N sodium sulfite
needed to dechlorinate the
rest of the biochemical
oxygen demand (BOD) sample.
2.	Measure the amount of
0.025 N sodium sulfite
just calculated.
3.	Add it to the entire BOD
sample.
4.	Stir or shake the sample
container so as to mix
the contents.
5.	Let the container stand
for 20 minutes.
la. Example calculation:
Assume 3.2 ml of the 0.025 N sodium sulfite were
used from the buret to titrate the 100 ml of
sample; i.e., 3.2 ml of the sodium sulfite were
needed to dechlorinate 100 ml. of sample.
lb. Assume 1 liter is the total volume of BOD sample.
1c. ml of 0.025 N sodium sulfite to dechlorinate the
rest of the BOD sample =
ml of entire
BOD sample (1000) ml of 0.025 N sodium
ml of BOD sample sulfite used from the
taken for the buret in C. (3.2)
above test (100)
= 1000 x 3 2
100 x
= 32
2a. It was 32 ml in the example calculation.
2b. Use a graduated pi pet if the required amount of
sodium sulfite is less than 10 ml. If it is
greater than 10 ml, use a graduated cylinder.





Page No. 3-15

-------
EFFLUENT MONITORING PROCEDURE: Dechlorination of Samples for Biochemical Ox.yqen Demand and
™Seeding of the Dilution Water
Page No. 3-16
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
E. Check on Dechlorina-
tion
1.	Repeat steps C.l through
C.7 above.
2.	Add 2 ml of starch
indicator.
3.	Swirl the flask to mix the
contents.
4.	Discard the 100 ml of
sample on which the check
was performed.
la. However, the color should not be red-brown at
this time. It will either be colorless, or
pale blue.
lb. Except use 100 ml of BOD sample to which the
0.025 N sodium sulfite solution has already been
added. (D.5.)
2a. Use a 10 ml graduated cylinder.
3a. If the solution is colorless, it indicates that
all of the chlorine has been "neutralized," and
the dechlorinated sample may now be used for the
BOD test. See the EMP on BOD5, Sections C.l.
and C.2.
3b. If there is an^ blue color, it indicates that not
all of the chlorine interferences have been
"neutralized."
3c. If there is blue color, add 2 drops of the 0.025 N
sodium sulfite to the BOD sample (1 liter in the
example calculation) and mix.
3d. Repeat 3c., E.l, E.2., and E.3., above until the
solution is colorless.
3e. When the colorless condition has been achieved,
the sample is dechlorinated and may be used for
the BOD test. See the Emp on BOD5, Sections C.l.
and C.2.

-------
rm iicmt wnnTTncTvr DDnrcnnDc. Dechlorination of Samples for biochemical Oxygen Demand and
EFFLUENT MONITORS PROCEDURE. Seeding Qf thfi Diluti£n Water
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
F. Calculation of Oxygen
Depletion in the
Sample, Due to Seed
1. Preparation of Seed
Controls
1. Pi pet 100 ml of the seed
(supernatant liquid) used
in B.1.2. into a 1 liter
graduated cylinder.
la. In order to calculate the BODc for a sample which
has been diluted with seeded dilution water, the
oxygen depletion in the sample due to the seed,
must be considered. In order to be useful for
the final calculation of BOD,-, the five-day
oxygen depletion in the seed control (F.l.) must
be 40-70%. In F.l. below, three seed dilutions
are set up; 10%, 15%, and 20%. At least one of
them should give a depletion in the 40-70% range.
If more than one dilution gives a depletion in
the 40-70% range, use the higher % value for cal-
culation of the BODr. The 10, 15, and 20% di-
lutions may not be nigh enough to give a 40-70%
depletion with some seed material. If not, ex-
perience will have to be used. For example, 20,
25, and 30% may have to be used. Three dilutions
should still be set up, however. The first step
in determining the oxygen depletion due to the
seed is to prepare seed controls. The second
step is to determine the oxygen depletion in the
seed controls. After this, a seed control cor-
rection is calculated and applied to the B0Dr
of the sample. Example calculations are useB
throughout the remainder of this procedure,
lb. Use a 100 ml volumetric pipet.

Page No. 3-17

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^ ..-..t -rorr.,^	Dech 1 orination of Samples for Biochemical Oxygen Demand and	p,np Mn o. 10
EFFLUENT MOMTORINO PROCEDURE: Seeding of the Diiution Water	Page N°" 3 18
OPERATING PROCEDURES
F. Calculation of Oxygen
Depletion in the
Sample. Due to Seed
(continued)
STEP SEQUENCE
2. Siphon dilution water (B.6.]
(20 C + 1°C) into the
graduated cylinder to the
1000 ml line.
3.	Thoroughly mix the contents
of the cylinder.
4.	Repeat steps 1., 2., and
3., except use 150 ml of
seed and a second cylinder,
5.	Repeat steps 1., 2., and
3., except use 200 ml of
seed and a third cylinder.
6.	Calculate the % of seed in
each of the 3 graduated
cylinders.
INFORMATION/OPERATING GOALS/SPECIFICATIONS
2a. If the siphon was "primed" with water, waste
about 50 ml before filling the cylinder.
2b. Cause no splashing of the liquid.
2c. Let the dilution water run down the sides of the
cylinder.
2d. Recall that dilution water is distilled water
plus the calcium, magnesium, ferric, and buffer
solutions.
3a. Use a plunger type mixer.
3b. Cause no splashing of the liquid.
4a. Use a 100 ml and a 50 ml volumetric pipet.
5a. Use a 100 ml volumetric pipet.
6a. 100 X 100 = 10 % of seed in the first cylinder.
1000
6b. 150 X 100
1000
6c. 200 X 100
1000
15% of seed in the second cylinder.
20% of seed in the third cylinder.
TRAINING
GUIDE NOTES

-------
EFFLUENT MONITORING PROCEDURE: Dechlorination of Samples for Biochemical Oxygen Demand and
Seeding of the Dilution Water
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
r. Calculation of Oxygen
Depletion in the
Sample, Due to Seed
(continued)
7.	Fill 2 BOD bottles from
the first (102) cylinder
by siphoning.
8.	Stopper the 2 BOD bottles
tightly and label them.
9.	Repeat steps 7 and 8 using
the second (15%) cylinder
and 2 more BOD bottles.
10.	Repeat steps 7 and 8 using
the third (20%) cylinder
and 2 more BOD bottles.
11.	Fill the flared top of 3
of the stoppered bottles
with water.
12.	Store them in the incubator
for 5 days.
7a. Hold the end of the siphon near the bottom of
the BOD bottle so as to prevent splashing.
7b. Open the siphon slowly.
7c. Fill the bottles until the liquid just begins
to overflow.
8a. Do not cause formation of an air bubble by
inserting the stopper too vigorously.
11a. line, of the 10%, one. Of the 15%, and fine, of the
20% bottles.
12a. At 20°C + 1°C.
12b. In the dark.
12c. Check the flared tops at least twice daily and
refill with water as needed.

Page No. 3-19

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EFFLUENT MONITORING PROCEDURE: Dechlorination of Samples for Biochemical Oxygen Demand and
~~————— Seeding of the Dilution Water
Page No. 3-20
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
F. Calculation of Oxygen
Depletion in the
Sample* Due to Seed
(continued)
2. Determination of
Oxygen Depletion
in the Seed Con-
trols
1
1.	Determine and record the
initial DO of the other
3 bottles.
2.	After five days, determine
and record the final DO of
of the 3 incubated bottles.
la. The other 10%, 15%, and 20% bottles,
lb. Use the Winkler Method - azide modification, or
a DO meter. See the appropriate effluent monitor'
ing procedure,
lc. The DO determination should be made within 15
minutes after mixing the contents of the liter
graduated cylinder.
2a. Use the same method as for the initial DO deter-
minations.
2b. Example calculation:
7.0 mg/1 = initial DO of the 20% seed control
2^8 mg/I = final DO of the 20% seed control
4.2 mg/1 = 5 day DO depletion of the 20% seed
control
4.2 X 100 = 60% DO depletion in the 20% seed
7.0 control. (This is within the 40-70%
range.)
By similar calculation, if the 15% seed control
gives a 40% depletion, use the 20% seed control
for calculating the final BOD,- value, as ex-
plained before.
2c. The 20% seed control will be used for the rest
of the example calculations.

-------
EFFLUENT MONITORING PROCEDURE:
Dechlorination of Samples for Biochemical Oxygen Demand and
Seeding of the Dilution Water
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
F. Calculation of Oxygen
Depletion in the
Sample, due to Seed
(continued)
3. Seeding of the
Dilution Water
Pipet the calculated
amount of seed (superna-
tant liquid) from the 2
liter flask (B.I.2.).
This is the same seed used
to prepare the seed con-
trols in F.l.
2.	Drain the seed from the
pi pet into the dilution
water container.
3.	Mix the dilution water and
seed by swirling the con-
tainer.
la
lb
For example, assume 3 liters of dilution water
are left after preparation of the seed controls
(F.l.).
Two ml of the supernatant liquid (seed) are used
for each 1 liter of dilution water - seed mixture,
lc. Continuing the example, 6 ml of seed would be
pipetted (3 x 2).
Id. As mentioned in the effluent monitoring procedure
on the Determination of Five-Day Biochemical
Oxygen Demand, during the incubation period, at
least 2 mg of dissolved oxygen/1 must be used by
the sample, and at least 1 mg/1 must remain at
the end of the incubation period. If less than
the 2 mg/1 is used, increase the amount of seed;
e.g., maybe 4 or 5 ml of seed will be needed for
each liter of the dilution water-seed mixture.
Only experience can determine the proper amount
of seed to use.
Use a graduated pipet to measure the seed. Ten
ml size in the example. If solids clog the tip
of the pipet, use a graduated cylinder. Ten ml
size in the example.
le
2a. Continuing the example, there are now 3006 ml of
liquid in the container.
3a. After mixing, the dilution water is said to be
seeded.
Page No. 3-21

-------
ccci iicnt mum t the TNT DDfir rni idc • Dechlorination of Samples for Biochemical Oxygen Demand and	page No. 3-22
EFFLUENT MONITORING PROCEDURE: Seeding of the Dilution Water
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
\ Calculation of Oxygen
Depletion in the
Sample, due to Seed
(continued)
4. Determination of
Seed Control Cor-
rection
4.	Place the container of
seeded dilution water in
the incubator.
5.	The 2 liter flask may now
be emptied.
1.	Determine what dilutions
will be made on the sample.
2.	Calculate the amount of
seeded dilution water
needed.
3.	Calculate the amount of
seed present in the di-
lution water used to
dilute the sample.
4a. 20°C + 1°C.
4b. Until it is needed.
la. See the effluent monitoring procedure on the
Determination of Five-Day Biochemical Oxygen
Demand, sections C.l.le. and C.1.If.
lb. For the purpose of example, assume you have
gained experience about the proper sample volume
to use, and you are going to set up a 40% di-
lution on the effluent of a treatment plant treat-
ing domestic wastewater.
2a. Continuing the example: the dilution is being
done in a 1 liter graduated cylinder.
2b. 1000 x 0.40 (40% as a decimal) = 400 ml sample.
2c. 1000 - 400 = 600 ml dilution water.
3a. Continuing the example:
600 ml of dilution water are being used.
3b. 6 ml of seed were added to 3 liters of dilution
water; i.e., 6 ml seed, and 3000 ml dilution
water.
^C"fi~+_300i5 X ^ = seec* in ttie seeclec:' dilution
water

-------
EFFLUENT MONITORING PROCEDURE:
Dechlorination of Samples for Biochemical Oxygen Demand and
Seeding of the Dilution Water
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
F. Calculations of Oxygen
Depletion in the
Sample. Due to Seed
(continued)
4. Calculate the % of seed
material in the 300 ml
BOD bottles containing
the sample.
5. Calculate the % of seed
in the seed control.
4a, Continuing the example:
A 40% dilution of the sample is being set up (as
was mentioned in F.4.1.1b.).
4b. In the 300 ml BOD bottle:
40% = sample
60% = seeded dilution water
4c. 300 X 0.4 (40% as a decimal) = 120 ml sample
300 X 0.6 (60% as a decimal) = 180 ml seeded
dilution water.
The seeded dilution water contains 0.2% seed
(F.4.3.3c. above).
180 X 0.002 (0.2% as a decimal) = 0.36 ml seed
material in the 300 ml BOD bottles.
4d. 0-36 x 100 = 0.12% seed in the BOD bottle
300
5a. Continuing the example:
The 20% seed control gave a 60% oxygen depletion
and will be used for the example calculation as
mentioned in F.2.2.2b. and F.2.2.2C.
5b. Therefore, there is 20% seed in the seed control.
Page No. 3-23

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EFFLUENT MONITORING PROCEDURE: Dechlorination of Samples for Biochemical Oxygen Demand and
			Seeding of the Dilution Water
Hage No. 3-24
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
F. Calculations of Oxygen
Depletion in the
Sample, Due to Seed
(continued)
6.	Calculate the seed control
correction factor.
7.	Calculate the seed cor-
rection.
6a. Continuing the example:
% seed in BOD bottle
% seed in the seed control
= 0.12
20
= 0.006
7a. Continuing the example:
seed correction = five day oxygen depletion in
seed control X factor.
7b. Oxygen depletion = 4.2 mg/1 (from F.2.2.2b.)
X 0.006
= 0.025 mg/1

G, Calculation of BOD,-,
Corrected for Use 8f
Seeded Dilution Water
1.	Calculate mg B0D,-/1 for
the sample.
2.	Subtract the seed correct-
ion.
la. See the effluent monitoring procedure on the
Determination of Five-Day Biochemical Oxygen
Demand, section D.l.
lb. Continuing the example:
Assume the B0D5 to be 30 mg/1.
2a. Continuing the example:
mg B0Dg/l = 30 mg/1 - 0.025 mq/1
= 29.98 mg/1
= 30 mg/1 (rounded off)

-------
EFFLUENT MONITORING PROCEDURE: Dechlorination of Samples for Biochemical Oxygen Demand and
" Seeding of the Dilution Water
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
G. Calculation of B0D5»
Corrected for Use
of Seeded Dilution
Water (continued)

2b. In this example, the seed correction was small
enough to be ignored. However, it will not
necessarily always be small enough to be ignored.





Page No. 3-25

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EFFLUENT MONITORING PROCEDURE: Dechlorination of Samples for Biochemical Oxygen
Demand and Seeding of the Dilution Water
TRAINING GUIDE
SECTION
TOPIC

I
Introduction

II
Educational Concepts
- Mathematics
III
Educational Concepts
- Science
IV
Educational Concepts
- Communications
V*
Field and Laboratory
Equipment
VI
Field and Laboratory
Reagents
VII
Field and Laboratory
Analysis
VIII
Safety

IX
Records and Reports

~Training guide materials are presented here under the headings marked *.
Page No. 3-26

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AFFLUENT MONITORING PROCEDURE: Dechlorination of Samples for Biochemical Oxygen Demand.
and Seeding of the Dilution Water
FIELD AND LABORATORY EQUIPMENT	Section V

TRAINING GUIDE NOTE
REFERENCES/RESOURCES
A. 1.1
If the glassware is especially dirty and cannot be
cleaned with ordinary detergents, chromic acid
cleaning may be required.


1.
Pour 35 ml of distilled water in a 250 ml beaker.


2.
Add about 1/8 teaspoon (simply estimate this
quantity) of sodium dlchromate, Na?Cr?07, to
the water.


3.
Swirl the beaker until the sodium dichromate
has dissolved.


4.
Keep repeating steps 2 and 3 until no more
sodium dichromate will dissolve.


5.
Pour the solution into a 2 liter beaker.


6.
Slowly pour 1 liter of concentrated sulfuric
acid, HgSO^, into the 2 liter beaker.
CAUTION: Use eyeglasses and protective clothing.


7.
Stir the mixture thoroughly.


8.
Store it in a glass stoppered bottle.


9.
The cleaning solution should be at a temperature
of about 50°C when It 1s used.


10.
It may therefore be necessary to warm the
cleaning solution.


11.
When using the warm cleaning solution, fill the
piece of glassware with the solution.


12.
Allow 1t to soak for 2-3 minutes (or longer).


13.
Pour the cleaning solution back into the storage
bottle.


14.
Rinse the piece of glassware ten times with tap
water.


15.
The cleaning solution may be reused until 1t
turns green.


16.
It should then be discarded.

Page No. 3-27

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A PROTOTYPE FOR DEVELOPMENT OF
ROUTINE OPERATIONAL PROCEDURES
for the
DETERMINATION OF DISSOLVED OXYGEN USING
A DISSOLVED OXYGEN METER
as applied in
WASTEWATER TREATMENT FACILITIES
and in the
MONITORING OF EFFLUENT WASTEWATERS
Developed by the
National Training and Operational Technology Center
Municipal Operations and Training Division
Office of Water Program Operations
U.S. Environmental Protection Agency
CH.O.do.EMP.lc.8.76
Page No. 4-1

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EFFLUENT MONITORING PROCEDURE: Determination of Dissolved Oxygen Using A
Dissolved Oxygen Meter
This process was developed by:
NAME Charles R. Feldmann
ADDRESS EPA, OWPO. NTOTC, Cincinnati, Ohio 45268
POSITION Chemist-Instructor
EDUCATION AND TECHNICAL BACKGROUND
B.S. - Chemistry
M.S. - Chemistry
1-1/2 years Industrial Chemist
4 years additional Graduate School
4 years college Chemistry Instructor
1-1/2 years DHEW - Air Pollution Program, Chemist
10 years DI - EPA, Chemist-Instructor
Page No. 4-3

-------
EFFLUENT MONITORING PROCEDURE-' Determination of Dissolved Oxygen Using A
Dissolved Oxygen Meter
1. Analysis Objectives:
The learner will use the attached EMP to place the Weston and Stack
Model 300 Dissolved Oxygen Meter into operation, including electrode
cleaning, membrane installation, calibration, and use of the meter to
make a dissolved oxygen measurement.
2.	Brief Description of Analysis:
The Winkler determination of dissolved oxygen-azide modification is
subject to many interferences. In the case of a BODg determination, the
problem is minimized to some extent because of sample dilution. If it
is felt, however, that appreciable amounts of interfering materials are
present, a dissolved oxygen meter should be used.
3.	Applicability of this Procedure:
At concentrations normally found in wastewater effluents, chlorine does
not affect the dissolved oxygen probe. Prolonged exposure to higher
concentrations of chlorine, and hydrogen sulfide, will necessitate cleaning
of the lead anode. Oil and grease will coat the membrane causing a de-
crease in sensitivity; the membrane should be replaced in this case.
This procedure was excerpted from the instruction book supplied with
the meter by the manufacturer.
Mention of a particular brand name does not constitute endorsement
by the U.S. Environmental Protection Agency
Page No. 4-4

-------
EFFLUENT MONITORING PROCEDURE: Determination of Dissolved Oxygen Using a
Dissolved Oxygen Meter
General Description of Equipment Used in a Process
A.	Capital
1.	Weston and Stack Model 300 Dissolved Oxygen (DO) Meter with Model
A-30 Probe, accessory kit and manufacturers instruction book
2.	Still, or other source of distilled water
3.	Trip balance, 100 g. capacity
B.	Reusable
1.	One 100 ml graduated cylinder
2.	One 250 ml Erlenmeyer Flask
3.	One 100 ml glass stoppered bottle
4.	One 200 ml plastic bottle
5.	One teaspoon
6.	Small blade screwdriver
7.	One 250 ml beaker
8.	Five ml syringe or eyedropper with tapered end
9.	Small pocket knife
10.	Four 300 ml BOD bottles
11.	Equipment for performing a Winkler DO determination-azide modification,
see the effluent monitoring procedure Winkler Determination of Dissolved
Oxygen-Az-ide Modification.
C.	Consumable
1.	Potassium iodide, KI, 50 g.
2.	Sodium sulfite, NagSO^, 25 g.
3.	Sodium hydroxide, NaOfl, 10 g.
4.	One rubber band
5.	Paper towels
6.	Silicone lubricant
7.	Source of distilled water
8.	One 1 inch long piece of scotch tape
9.	Reagents for performing a Winkler DO determination-azide modification,
see the effluent monitoring procedure Winkler Determination of Dissolved
Oxygen-Azide Modification.
10.	Sodium dichromate, Na-jCr^
11.	Concentrated sulfuric acid, H2SO4
12.	Soap
(Items 10, 11, and 12 are for cleaning glassware. The quantities
will therefore vary.)
13.	Brushes (for cleaning glassware)
14.	Brush (for cleaning balance)
15.	Sponges (for cleaning of laboratory table tops)
Page No.

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EFFLUENT MONITORING PROCEDURE: Determination of Dissolved Oxygen Using a
a Dissolved Oxygen Meter
C. Continued
All reagents should be of high quality. Different chemical
manufacturers may have different ways of indicating a high quality
reagent. While no endorsement of one chemical manufacturer over
another 1s intended, the following are some designations used in
four chemical catalogs to indicate high quality reagents.
Catalog	Designations
Thomas	Reagent, ACS, Chemically
Matheson, Coleman & Bell
Curtin Matheson
Scientific, Inc.
Pure (CP)
Reagent, ACS
Primary Standard, ACS, AR
Fisher
Certified, ACS
Page No. 4-6

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FRONT VIEW OF METER
meter scales -
(HI, LO, temp)
on off/chg
switch
DO probe
attachment
function switch
zero check switch
temp test button
cal
zero *
Paqe Mo. 4-7

-------
REAR VIEW OF METER
tubes for
accessory
storage
0 probe attachment
temp test button
!!
AC power cord attachment
Page No. 4-8

-------
CUT-AWAY VIEW OF PROBE, SHELL, AND STIRRING MECHANISM
service cap
0" rings
cord to meter,
electrolyte filling
screw (and washer)
battery box
long screw
PROBE
anode rim
thermistor pins in well
lead coil anode"
holes
(two)
flange
nipple and "0" ring/
(seats in thermistor
pin well)
spring clip
platinum cathode
SHELL
micro switch
micro switch actuator arm
shield (to protect
membrane)
Page No.4-9

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EFFLUENT MONITORING PROCEDURE: Determination of Dissolved Oxygen Using a Dissolved Oxygen Meter
Page No. 4-10



TRAINING
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
GUIDE NOTES
A. Equipment Preparation



1. Glassware
1. Clean all glassware and

V.A.I.1

rinse with distilled water.

(p. 22}
2. Balance inspection
1. Check all balances for



cleanliness and proper



operation.


B. Reagent Preparation



1. Electrolyte
1. Weigh 50 g. of potassium


solution
iodide, KI.



2. Weigh 0.1 g. of sodium



sulfite.



3. Dissolve the two solids in
3a. Unless otherwise specified, the term water


100 ml of water.
means distilled water.



3b. Unless otherwise specified, solutions should be



stored in glass stoppered bottles.


4. Store the electrolyte in a



small bottle.


2. Sodium hydroxide
1. Weigh 10 g. of sodium


solution
hydroxide, NaOH.



2. Dissolve it in 90 ml of



water.



3. Store the solution in a



small plastic bottle.


3. Sodium sulfite
1. Measure 1 teaspoon of sodium


solution
sulfite, Na2S02>



2, Dissolve it in 500 ml of
2a. Prepare this solution just prior to use.


tap water.

-------
t>H.Ut.NI MONITORING PROCEDURE: Determination of Dissolved Oxygen Using a Dissolved Oxygen Meter
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
Equipment Preparation
1. Battery check -
Weston and Stack
Model 300 Dissolved
Oxygen (DO) Meter
2. Battery check-DO
probe stirring
mechanism
1. Check the power cord attachec
to the rear of the meter.
2.	Turn the function switch to
the temperature position.
3.	While pressing the temp test
button (left side of meter),
adjust the temp adj screw
(right side of meter) to
read 50°C (bottom scale on
front of meter).
4.	Turn the function switch to
the transit position.
1.	Remove the long screw from
the top of the battery box.
2.	Remove the top half of the
battery box.
3.	Insert two size AA 1-1/2
volt batteries (provided with
the instrument) into the
battery box.
4.	Place the top on the battery
box.
5.	Insert the long screw into
the top of the battery box.
6.	Screw it down.
7.	Using your finger, close the
micro switch actuator arm on
the side of the probe.
la. The meter is portable and the cord should be
plugged in to recharge the nickel cadmium
battery during storage periods.
3a. One should be upright, the other upside down.
4a. Note that there is a tip and a hole on the bottom
of the top half of the battery box. These fit in>
to a hole and a tip on the top of the bottom half
of the battery box.
7a. The probe stirring mechanism will start.
Page No. 4-11

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EFFLUENT MONITORING PROCEDURE: Determination of Dissolved Oxygen Using a Dissolved Oxygen Meter
Paae No. 4-12
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
2. Continued
3. Cathode check-DO
probe
4. Anode check-DO
probe
8.	Loosen the long screw,
9.	Raise the top half of the
battery box about half inch.
10. Wrap a rubber band around
the battery box.
10a. The rubber band should be placed in such a way
that the two halves of the battery box are kept
apart.
10b. This will keep the stirring mechanism from
operating when the probe is not in use.
10c. Do not remove the rubber band and tighten the
long screw until the meter is to be calibrated
or measurements are to be made.
1.	Unfasten the spring clip and
remove the probe from the
shell.
2.	Examine the platinum cathode
at the end of the probe.
1.	Remove the service cap on
top of .the probe.
2.	Examine the two black "0"
rings.
3.	Coat the two "0" rings with
a very thin layer of silicone
lubricant.
4.	Invert the probe over a
table top.
2a. It should be free of dirt.
2b. If it is not, wipe it briskly with a paper towel,
or coarse piece of cloth.
2a. They should be free of dirt.
4a. The lead coil anode should drop out.
4b. If it does not, tap the probe lightly on the table
top.

-------
EFFLUENT MONITORING PROCEDURE: Determination of Dissolved Oxygen Using a Dissolved Oxygen Meter
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
4. Continued
5.
Examine the anode.
5a.
It should be free of dirt and corrosion. Yellow
colored corrosion is common.


6.
If corrosion is present, soak
the anode in the sodium
hydroxide solution.
6a.
6b.
A few minutes soaking should suffice.
Very minute amounts of corrosion will not
cause problems.


7.
Rinse the anode thoroughly
with tap water.




8.
Rinse the anode thoroughly
with distilled water.




9.
Examine the rim, inside of
the probe, on which the
anode sits.
9a.
It should be free of dirt and corrosion. Yellow
colored corrosion is common.


10.
If corrosion is present,
scrape it away using the
blade of a small screwdriver.
10a.
A swab dipped in the sodium hydroxide solution
will also remove the corrosion.


11.
Rinse the rim and interior of
the probe thoroughly with tap
water and then with distilled
water.



5. Thermistor contact
check
1.
2.
Invert the probe.
Examine the two thermistor
pins in the small well.
2a.
2b.
They should be free of dirt and corrosion.
If they are corroded, gently scrape them, using
the blade of a small screwdriver.


3.
Examine the "0" ring around
the nipple which seats in the
thermistor pin well.
3a.
It should be free of dirt.

Page No. 4-13

-------
EFFLUENT MONITORING PROCEDURE: Determination of Dissolved Oxygen Using a Dissolved Oxygen Meter
Page No. 4-14
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
5.	Continued
6.	Membrane
Installation
4. Coat it with a very thin
layer of silicone lubricant.
1.	Select a 1 mil membrane
(furnished with the
instrument).
2.	Examine it in bright light
for holes.
3.	Hold the probe upside down.
4.	Lay the square membrane over
the platinum cathode.
5.	Fold the membrane back over
the probe.
6.	Loop a small rubber band
(furnished with the instru-
ment) around the probe three
times just above the flange.
7.	Gently pull on the loose
edges of the membrane.
8.	Place a short piece of scotch
tape over the well containing
the two thermistor pins.
9.	Remove the electrolyte fill-
ing screw and washer.
10.	Hold the probe in a vertical
position, with a finger tip
held over the electrolyte
la. A mil is 0.001 inch.
lb. One-half mil membranes are sometimes used. They
respond faster, but are more fragile.
2a. If any are seen, discard the membrane.
4a. The cathode should be in the center of the square.
6a. The rubber band should hold the membrane snugly,
but not too tightly.
6b. Two turns of the rubber band may suffice in some
cases.
7a. There should be no folds in the membrane over the
platinum cathode.
7b. The pulling should not, however, cause tearing
of the membrane.
8a. This will prevent moisture from getting into the
thermistor pin well while the probe is being
filled with electrolyte.
10a. The cathode should point down.

-------
EFFLUENT MONITORING PROCEDURE: Determination of Dissolved Oxygen Using a Dissolved Oxygen Meter
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
6. Continued
11.	Pour electrolyte solution
into the service cap opening.
12.	With a twisting motion, slide
the rubber band and membrane
down about 1/2 inch.
13.	With a similar motion, slide
the rubber band and membrane
back up to its original
position.
14.	Loop a second rubber band
around the membrane about
half-way between the first
rubber band and the two
holes near the platinum
cathode.
15.	Carefully drop the lead anode
into place.
16.	Screw in the service cap
about half-way.
17.	Hold the probe in a hori-
zontal direction, electrolyte
filling hole up.
18.	Remove the finger tip from
the electrolyte filling well.
19.	Using a syringe or eyedropper
with a tapered end, add
additional electrolyte
through the filling hole.
11a. Fill the probe interior almost to the top.
12a. The pocket formed by the membrane will fill with
electrolyte.
13a. There should be no wrinkles in the part of the
membrane lying across the platinum cathode.
14a. Wrap the rubber band as tightly as possible, one
turn next to another.
14b. There should still be no wrinkles in the part of
the membrane lying across the platinum cathode.
14c. It may be awkward to keep a finger tip over the
electrolyte filling well.
14d. No problem is created if some electrolyte is
lost, it will be replaced later.
18a. Some electrolyte will probably run out.
19a. Gently rock the probe back and forth after each
addition so as to dislodge air bubbles.
19b. Tap the sides of the probe with a pen or pencil
to ensure bubble escape.

Page No. 4-15

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EFFLUENT MONITORING PROCEDURE: Determination of Dissolved Oxygen Using a Dissolved Oxygen Meter
Page No. 4-15
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
6. Continued
20.	Tighten the service cap.
21.	Check for completeness of
electrolyte filling.
22.	Replace the electrolyte
filling screw and washer.
23.	Ensure that no air bubbles
have formed under the mem-
brane covering the platinum
cathode.
24.	Cut the membrane around the
probe between the two rubber
bands.
25.	Cut away the first rubber
band.
26.	Remove the excess membrane.
27.	Carefully rinse the outside
of the probe and membrane
with water.
28.	Gently shake off the water.
29.	Remove the piece of tape
covering the thermistor pin
wel 1.
30.	Carefully place the probe
back in the shell.
31.	Place the probe in a 300 ml
BOD bottle full of water.
23a. If there are any, the membrane should be removed
and reinstalled.
30a. Be cautious not to tear the membrane.
30b. Make sure the spring clip is properly closed.

-------
EFFLUENT MONITORING PROCEDURE: Determination of Dissolved Oxygen Using a Dissolved Oxygen Meter
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
6. Continued
7. DO meter zeroing
32. With your eyes at the same
level as the end of the
probe, look carefully at the
end of the probe for about
one minute.
1. Attach the DO probe to the
DO meter.
2.	Rinse the outside of the
shell with water.
3.	Place the probe in a 300 ml
BOD bottle filled with
sulfite solution.
4.	Allow it to stand for
10 minutes.
5.	Remove the rubber band from
the stirring mechanism
battery box.
6.	Screw down the long screw
in the top of the battery
box.
7.	Wait two minutes.
8.	Turn the on off/chg toggle
switch to the on position.
9.	Turn the function switch
to the HI mg/liter position.
10. When the needle reads 1.5 on
the top scale, turn the func-
tion switch to the LO mg/liter
position.
32a. If there are any holes in the membrane the
leaking electrolyte will be seen in the water,
and the membrane must be replaced.
la. A pliers may be used to assure a snug fit, but
be careful not to damage the knurls on the
locking collar.
2a. Do not get water on the micro switch of the
stirring mechanism.
6a. The stirring mechanism should start since the
flared top of the BOD bottle closes the micro
switch actuator arm.
9a. The needle on the meter face will move to the
right and then slowly drift to the left.
10a. The needle should continue to drift to the left.
Page No. 4-17

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EFFLUENT MONITORING PROCEDURE: Determination of Dissolved Oxygen Using a Dissolved Oxygen Meter
Page No. 4-18
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
7. "Continued
8. DO meter
calibration
11.	Wait one additional minute.
12.	Depress the zero check toggle
switch and turn the zero
screw (bottom front of the
instrument) until the needle
reads 0 on the middle scale
of the meter.
13.	Turn the on off/chg toggle
switch to the off/chg
position.
4.	Turn the function switch
to the transit position.
5.	Loosen the 7ong screw and
raise the top half of the
battery box.
6. Replace the rubber band which
separates the two halves of
the battery box.
17.	Remove the probe from the
BOD bottle.
18.	Rinse off the bottom of the
probe thoroughly.
19.	Place the probe in a BOD
bottle filled with water.
1. Fill two 300 ml BOD bottles
to overflowing with distilled
water.
12a. After releasing the toggle switch the needle
may slowly drift toward a "true" zero.
13a. In the off/chg position, the nickle cadmium
battery is charging when the power cord is
attached.
14a. For the remainder of this EKP, this procedure
will be referred to as turning the stirring
mechanism off. Lowering the top half of the
battery box and tightening the long screw will
be referred to as turning the stirring mechanism
on.
17a. Be cautious not to get the micro switch wet.
17b. All traces of the sulfite solution must be
removed.
18a. To prevent the membrane from drying out, always
keep the probe in water when not in use.
la. It is essential that the two samples be identical
in oxygen content.

-------
EFFLUENT MONITORING PROCEDURE: Determination of Dissolved Oxygen Using a Dissolved Oxygen Meter
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
8. Continued
2.	Determine the DO of one of
the bottles.
3.	Remove the probe from the BOD
bottle containing the water.
4.	Rinse the end of the probe
with distilled water.
5.	Place the probe in the second
BOD bottle filled with dis-
tilled water.
6.	Turn the on off/chg toggle
switch to the on position.
7.	Turn the stirring mechanism
on.
8.	Turn the function switch to
the HI mg/liter position.
9.	Wait about two minutes.
!0. Turn the cal screw (bottom
front of the meter) until the
needle reads (on the upper
scale) the same DO value as
was obtained from the
Winkler titration.
11. Turn the stirring mechanism
off.
2a. Use the Winkler Method-Azide Modification.
9a. Because the slower responding 1 mil membrane is
being used, about two minutes should be required
for the needle reading to stabilize.
9b. With some meters, experience may indicate that a
shorter waiting period will suffice.

Page No. 4-]9

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EFFLUENT MONITORING PROCEDURE: Determination of Dissolved Oxygen Using a Dissolved Oxygen Meter
Page No. 4-;'G
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
8. Continued
12.	Turn the on off/chg toggle
switch to the off/chg
position.
13.	Turn the function switch
to the transit position.
14.	Place the probe back in the
BOD bottle of distilled
water. The membrane should
always be kept wet when not
in use.
13a. There is no firm rule about how often to zero
and calibrate the Weston and Stack Model 300
DO meter.
13b. A conservative estimate would be to do the
calibration daily, and the zeroing every other
day in times of frequent use.
13c. Both steps should be performed if the meter has
not been used for several days.
13d. After installation of a new membrane, the
calibration changes markedly during the first
24 hours, and frequent calibrations are needed
during this period.
I
1

-------
TRAINING GUIDE
SECTION	TOPIC
I	Introduction
II	Educational Concepts - Mathematics
III	Educational Concepts - Science
IV	Educational Concepts - Communications
V*	Field & Laboratory Equipment
VI	Field & Laboratory Reagents
VII	Field & Laboratory Analysis
VIII	Safety
IX	Records & Reports
*Training guide materials are presented here under the headings marked *.
Page No. 4-21

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EFFLUENT MONITORING PROCEDURE: Determination of Dissolved Oxygen Using a
Dissolved Oxygen Meier
TRAIIIiriG GUIDE MOTE
If the glassware is especially dirty and Cci"n.>l be
cleaned with ordinary detergents, chromic jciJ
cleaning may be required.
1.	Pour 35 ml of distilled water in a 260 ml boaker.
2.	Add about 1/8 teaspoon (simply estimate this
quantity) of sodium dicrhomate, Na?Cr0H7
to the water.
3.	Swirl the beaker until the sodium dichromate
has dissolved.
4.	Keep repeating steps 2 and 3 until no more
sodium dichromate will dissolve.
5.	Pour the solution into a 2 liter beaker.
6.	Slowly pour 1 liter of concentrated sulfuric
acid, H2SO4, into the 2 liter beaker.
Caution: Use eyeglasses and
protective clothing.
7.	Stir the mixture throughly.
8.	Store it in a glass stoppered bottle.
9.	The cleaning solution should be at a temperature
of about 50°C when it is used.
10.	It may therefore be necessary to warm the
cleaning solution.
11.	When using the warm cleaning solution, fill the
piece of glassware with the solution.
12.	Allow it to soak for 2-3 minutes (or longer).
13.	Pour the cleaning solution back into the storage
bottle.
14.	Rinse the piece of glassware ten times with
tap water.
15.	The cleaning solution n.ay be reused until it
turns green.
16.	It should then be discarded.
Section V
REFERCNCES/RESOURC!
13th Standard Methods,
p. 135, section 2.C.2
Page No. 4-22

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A PROTOTYPE FOR DEVELOPMENT OF
ROUTINE OPERATIONAL PROCEDURES
for the
DETERMINATION OF DISSOLVED OXYGEN IN
WASTEWATER: POLAROGRAPHIC PROBE METHOD
as applied in
WASTEWATER TREATMENT FACILITIES
and in the
MONITORING OF EFFLUENT WASTEWATERS
Developed by the
National Training and Operational Technology Center
Municipal Operations and Training Division
Office of Water Program Operations
U.S. Environmental Protection Agency
CH.O.do.EMP.3.3.77
Page No. 5-1

-------
EFFLUENT MOiilTURING PROCEDURE: Determination of Dissolved Oxygen:
Polarographic Probe Method (YSI Model
54 Oxygen Meter)
This instructional sequence was prepared by:
NAME Paul F. Hall bach
ADDRESS EPA, OWPO, NTOTC, Cincinnati, Ohio 45268
POSITION Chemist-Instructor
EDUCATION & TECHNICAL BACKGROUND
8S - Chemistry
14 years Industrial Chemist
16 years HEW-FWPCA-EPA-Chemist
Page No. 5-3

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FFFLUENT MONITORING PROCEDURE: Determination of Dissolved Oxygen:
Polarographic Probe Method (YSI Model
54 Oxygen Meter)
1.	Analysis Objectives:
The operator will be able to set up, calibrate and use a YSI oxygen
meter for the determination of dissolved oxygen in a sample of waste-
water treatment plant effluent.
2.	Brief Description of Analysis*:
The meter is set up and calibrated and the polarographic probe is
inserted into the appropriate sample. A reading is obtained from the
meter which correlates the dissolved oxygen concentration in the
sample.
~Standard Methods for the fxamination of Water and Wastewater, I4t.h Ld.,
1975. APHA, Washington, DC, p. 4b()
Page #6. 5-4

-------
EFFLUENT MONITORING PROCEDURE: Determination of Dissolved Oxygen:
Polarographic Probe Method (YSI Model
54 Oxygen Meter)
General Description of Equipment Used in the Process
A.	Capital Equipment
1. Dissolved oxygen meter and polarographic probe assembly - Yellow
Springs Instrument Company
B.	Reusable
1.	B.O.D. bottle (300 ml)
2.	One plastic squeeze bottle
3.	Eyedropper bottle
4.	Scissors
5.	Small screwdriver
C.	Consumable
1.	Standard membranes (0.001" - YSI #5352)
2.	Probe Service Kit (YSI #5034)
Page No. 5-5

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AFFLUENT MONITORING PROCEDURE: Determination of Dissolved Oxygen in Wastewater- Pole.rographic Proie Method
?aqp No. 5-s
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
A. Probe Preparation
1.	Add distilled water to the
KC1 crystals and dissolve
completely.
2.	Transfer a part of the KC1
solution to the eyedropper
bottle.
3.	Remove the protective mem-
brane and "0" ring.
4.	Select a membrane from the
membrane packet.
b. Support the probe in a
vertical position.
6.	With, one thumb secure one
end of the membrane to the
side of the probe.
7.	With the eyedropper, fill
the central hole avoiding
air bubbles.
8.	Wet the gold electrode
and the lucite around it.
la. KC1 crystals included in YSI Probe Service Kit
(Part no. 5034) a saturated KC1 solution diluted
1:1 with distilled water should be used.
4a. Lay on a clean sheet of paper. Handle only by
the ends.
4b. Use only YSI recommended membranes and filling
solution. Distilled water must be used in
making the KC1 solution. Tap water contains
iron and other salts that result in poor
electrode performance and will contaminate
the electrodes and result in snort life.
8a. The surface tension of the KC1 will cause a
large drop or meniscus to form above the
electrode. This will ensure complete contact
between the membrane a.nd the KC1.

-------
EFFLUENT HONITORING PROCEDURE: Determination of Dissolved Oxygen in Wastewater: Polarographic Probe Hethod
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
A. Continued
9. Stretch the membrane over
the top of the electrode.
10.	Stretch an- "0" ring into
place—inspect for winkle-
free membrane.
11.	Remove the excess membrane
about 1/8" beyond the "0"
ring with scissors.
12.	A small air bubble may
appear under the membrane.
13.	After the air has been
driven from the anode,
remove the membrane, refill
with KC1, and install
another membrane.
14.	Rinse probe with distilled
water.
15.	The probe 1s ready for
operation.
10a. A taut smooth membrane surface is required. A
lax membrane will result in erratic performance,
slow speed of response and poor shock performance.
12a. This is normal, however, strive for a bubble-free
probe. New probes, or probes that have been
allowed to dry out, will continue to develop
bubbles until the porous anode is completely
filled.

B. Calibration
1.	Connect the two probe plugs
to the jacks on the side of
the instrunent.
2.	VJ1th the instrument turned
off check the mechanical
zero of the meter—pointer
should indicate zero.
2a. Adjust with the screw on the meter case. Recheck
y/hen the position of instrument is changed.

Page No. 5-7

-------
EFFLUENT MONITOR!'.3 PROCEDURE: Determination of Dissolved Oxygen in Wastewater: Polarographic Probe Method
Daqe No. 3-S
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE N0TES
6. Continued
3.	Switch to the RED LINE
position and adjust the
meter to red line with the
front panel control.
4.	Place the probe in the BOD
bottle containing a water
sample of known dissolved
oxygen content.
5.	Turn the stirring mechanism
switch on.
6.	Switch to the TEMP position
and read the temperature
when the meter is steady.
7.	Switch to the ZERO position
and adjust the meter to zero
with the ZERO control.
8.	Switch to the 0-10 ppm
position and calibrate the
instrument with the CAL
control.
9.	Turn off the stirring
mechanism and the instrument.
4a. Samples of known oxygen concentration can be ob-
tained by analyzing a duplicate sample by the
Winkler Titration Method.
8a. Calibration and measurement should be carried out
on the same range to avoid compounding meter
tolerance error.

C. Dissolved Oxygen
Measurement
1.	Place the probe in the BOD
bottle containing the un-
known sample.
2.	Turn on the stirring mechan-
ism and the instrument.

i
!

-------
EFFLUENT MONITORING PROCEDURE: Determination of Dissolved Oxygen in Wastewater: Polarographic Probe Method
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATICNS
training
GUIDE NOTES
C. Continued
3.	Switch to the 0-10 ppm
position and read the
dissolved oxygen concen-
tration obtained.
4.	Turn off the stirring
mechanism and the instrument.


Page No. b-9

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A PROTOTYPE FOR DEVELOPMENT OF
ROUTINE OPERATIONAL PROCEDURES
for the
pH DETERMINATION OF WASTEWATER AND WASTEWATER TREATMENT
PLANT EFFLUENTS
as applied 1n
WASTEWATER TREATMENT FACILITIES
and in the
MONITORING OF EFFLUENT WASTEWATERS
Developed by the
National Training and Operational Technology Center
Municipal Operations and Training Division
Office of Water Program Operations
U.S. Environmental Protection Aqencv
CH.pH.EMP.1.3.77
Page No. 6-1

-------
EFFLUENT MONITORING PROCEDURE: Measurement of pH
This Instructional sequence was developed by:
NAME Paul F. Hall bach
ADDRESS EPA. OWPO, NTOTC, Cincinnati, Ohio 45268
POSITION Chemist Instructor
EDUCATION AND TECHNICAL BACKGROUND
B.S. Chemistry
14 years Industrial Chemist
16 years HEW-FWPCA-EPA-Chemist

-------
EFFLUENT MONITORING PROCEDURE: Measurement of pH
1.	Analysis Objectives:
WWTP operator will set up, calibrate and operate portable type pH meter for the
pH measurement of wastewater and WWTP effluent.
2.	Brief Description of Analysis*
A portable type, battery operated pH meter, equipped with a glass electrode
system is used to measure the pH of wastewater treatment plant samples.
3.	Applicability of this Procedure:
a.	Range of concentrations:
pH scale 0-14
b.	Pretreatment of Sample:
None
c.	Treatment of interferences in samples:
None
*Standard Methods for the Examination of Water and Wastewater, 14th Ed., 1975, APHA,
Washington, D.C., p. 460
Page No. 6-4

-------
EFFLUENT MONITORING PROCEDURE : Measurement of pH
General Description of Equipment Used 1n the Process
A.	Capital Equipment
1. pH Meter IL Model 175 PORTO-matic*
The IL Model 175 PORTO-matic pH meter is a small, solid state, battery
operated, portable Instrument for the measurement of the pH of aqueous
solutions. Manufacturer's Specifications are as follows:
pH range: 0-14
pH Scale: 7.2", 1/2% zero centered
Readability: 0.01 pH
Electrical Accuracy: better than 0.035 pH
Drift per Day: less than 0.01 pH
Battery Life: 2000 hours
B.	Reusable
1.	Wash bottle, plastic
2.	Beakers, 250 ml, 150 ml, 25 ml
C.	Consumable
1.	Buffer Solution pH 4
2.	Buffer Solution pH 9
3.	Buffer Solution pH 6.9
4.	Buffer Solution pH 7.4
5.	Saturated KC1 Solution
*Ment1on of a specific brand name does not constitute endorsement by the
U.S. Environmental Protection Agency
Page No. 6-5

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EQUIPMENT - PORTO-MATIC pH METER
OPERATING CONTROLS FRONT PANEL, 175 PORTO-MATIC pH METER
—————	1	
HANDLE
METER -
SCALE pH 0-14
PORTO-MATIC
pH METER
MODEL 175
TEMPERATURE
COMPENSATION
CONTROL
POWER SWITCH
- OFF/ON	
BALANCE
KNOB"^
40. ,60
20
80
BALANCE
FIGURE 1
REAR PANEL, 175 PORTO-MATIC pH METER
pH ELECTRODE
SPRING CLIP
INPUT
CONNECTOR
PLASTIC ^
ELECTRODE CAP
LOCK NUTS
PLASTIC
' CASE
FIGURE 2
Page No. 6-7

-------
EFFLUENT MONITORING PROCEDURE: pH Determination of Wastewater and
Wastewater Treatment Plant Effluents
Page No. 6-0
OPERATING PROCEDURES
STEP SEQUENCE
INF0RMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
A. Instrument Setup
1.	Place pH meter on solid
surface.
2.	Remove electrode from rear
panel.
3.	Twist the cap at the bottom
of the electrode to align
the filling holes.
4.	Check the level and satura-
tion of the KC1 solution in
the reference chamber of the
electrode.
5.	Place the electrode in a
150 ml beaker containing
100 ml of distilled water.
6.	Turn ON/OFF switch on.
See Fig. 2.
See Fig. 2.
I.A
(p. 13)
B. Meter Calibration
1.	Set temperature compensation
knob to correspond to tem-
perature of buffer solution
to be used for calibration.
2.	Transfer 50 ml of pH 6.9
buffer solution into a
clean 150 ml beaker.
3.	Turn meter switch "off"
la. Previously prepared standard buffer solution pH
6.9 should be used. Buffer solutions can be pre-
pared from the formulas shown in the attached
table.
3a. Meter should be "off" when electrode is out of
solution.
VII.1
(P. 15)

-------
EFFLUENT MONITORING PROCEDURE: pH Determination of Wastewater and
"	Wastewater Treatment Plant Effluents
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES

4.	Rinse the electrode with
buffer solution and immerse
it in the beaker containing
the pH 6.9 buffer.
5.	Turn meter on.
6.	Adjust the needle on the
meter to read 6.9 by turning
the balance knob clockwise
or counter-clockwise.
7.	Turn the power switch off.
8.	Repeat calibration with
buffer pH 7.4
9.	Remove the electrode from
the buffer and rinse the
electrode three times with
distilled water.
10.	Add distilled water to cap.
11.	Twist the cap on the bottom
of the electrode so that the
filling holes are closed
and water surrounds the
glass membrane.
12.	Discard buffer solution.
4a. Do not allow bubbles to collect around the
ceramic junction of the reference chamber.
6a. Allow adequate time for the glass electrode to
come into equilibrium with the sample (approxi-
mately 30 seconds).
9a. Use a squeeze type wash bottle.
11a. The pH sensitive membrane dehydrates when removed
from water. Dry glass electrodes should be soaked
in buffer or water for several hours before use.
12a. Never pour used buffer solution back into buffer
bottle.
VII.1
{p. 15)




Page No. 6-9

-------
EFFLUEifl" KG?iIT0Rli<3 PROCEDURE: pH Determination of Wastewater and
Wastewater Treatment Plant Effluents
Paae No. 6-10
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GLIDE NOTES
C. Use of Instrument for
pH measurement
1.	Adjust temperature compen-
sation knob to the tempera-
ture of the unknown solution.
2.	Twist open the electrode cap.
3.	Immerse the electrode into
the unknown.
4.	Turn the power switch on.
5.	Allow adequate time for the
glass electrode to come into
equilibrium with the sample
(approximately 30 seconds).
6.	Determine pH of unknown
solution by observation of
meter needle on pH scale
of instrument.
7.	Enter the result on the
appropriate report form.
Record your value to the
nearest 0.1 pH unit.
8.	Turn off instrument.
9.	Rinse the electrode with
distilled water.
10.	Add water or buffer to cap
prior to closing to prevent
dehydration of electrode.
11.	Close the cap of the
electrode.
5a. Do not allow bubbles to collect around the
ceramic junction of the reference chamber.
6a. Swirl probe several times before taking reading.
6b. Take reading with mirror reflection of needle
obscured by needle.

-------
EFFLUENT MONITORING PROCEDURE: pH Determination of Wastewater and
Wastewater Treatment Plant Effluents
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
D. Maintenance
1.	Check the level and satura-
tion of potassium chloride
in the reference chamber of
electrode.
2.	Keep the glass membrane wet
with distilled water when
not being used.
3.	Do not contaminate standard
buffer solution.
4.	Turn the instrument off
when not 1n use.
la. The reference chamber of the pH electrode system
should always be kept nearly full with saturated
potassium chloride solution.
4a. The pH meter is a battery-operated instrument.
VII.D
(p. 15)
E. Trouble Shooting
1.	Erratic needle movement:
a.	Fill the measuring
chamber completely
b.	Soak the external surface
of the ceramic plug in
warm water.
c.	Resaturate the reference
chamber.
2.	No instrumental response
when measurement is taken.
la. Erratic needle movement:
Can be caused by bubbles around the ceramic
reference junction.
Can be caused by contamination or salt
crystallization of the reference ceramic
plug.
Can be caused by unsaturation of the reference
chamber.
2a. Exchange electrode or electrodes with new
electrodes.
2b. If porous plug of electrode is clogged, take
appropriate action to clean. See electrode
specification sheet.

Page No. 6-11

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EFFLUENT MONITORING PROCEDURE: pH Determination of Wastewater and
Wastewater Treatment Plant Effluents
TRAINING GUIDE
SECTION	TOPIC
I*	Introduction
II	Educational Concepts - Mathematics
III	Educational Concepts - Science
IV	Educational Concepts - Communications
V	Field and Laboratory Equipment
VI	Field and Laboratory Reagents
VII*	Field and Laboratory Analysis
VIII	Safety
IX	Records and Reports
~Training guide materials are presented here under the heading marked *,
These standardized headings are used throughout this series of procedures.
Dane No. 6-12

-------
EFFl.UF.HT MONITORING PROCEDURE: pH Determination of Wastewater and
Wastewater Treatment Plant Effluents
SECTION I
TRAINING GUIDE NOTE	REFERENCES/RESOURCES
I	Introduction
1. pH General Considerations
pH is a term used to describe the in- Nebergall, W. H., Schmidt, F. C. and
tensity of the acid or alkaline condition Holtzclaw, Jr., HF., College Chem.,
of a solution. The concept of pH evolved 2nd Ed. Heath & Co., Boston, 1963
from a series of developments that led to
a fuller understanding of acids and alka-
line solutions (bases). Acids and bases
were originally distinguished by their
difference in physical characteristics
(acids-sour, bases-soapy feel). In the
18th century it was recognized that adds
have a sour taste (vinegar-acetic acid),
that they react with limestone with the
liberation of a gaseous substance (carbon
dioxide) and that neutral substances
result from their interaction with alka-
line solutions.
Acids are also described as compounds
that yield hydrogen ions when dissolved
in water. And that bases yield hydroxide
ions when dissolved in water. The pro-
cess of neutralization is then considered
to be the union of hydrogen (H+) ions and
hydroxy! (OH") ions to form neutral water
(H+ + OH—*H20).
It has been determined that there are
1/10,000,000 grams of hydrogen Ions and
17/10,000,000 grams of hydroxyl ions inone
liter of pure water. The product of the
H+ and OH- 1ons equal a constant value.
Therefore, if the concentration of the H+
ions 1s Increased there is a corresponding
decrease in OH" ions. The acidity or
alkalinity, hydrogen 1on concentration
of a solution 1s given 1n terms of pH.
The pH scale extends from 0 to 14 with
the neutral point at 7.0.
Page Ho. 6-13

-------
EFFI.UI.NT MONITORING PROCEDURE: pH Determination of Wastewater and
Wastewater Treatment Plant Effluents
TRAINING GUIDE MOTE
REFERENCES/RESOURCES
2. Electrode Design
About 1925 it was discovered that an
electrode could be constructed of glass
which would develop a potential related
to the hydrogen-ion concentration with-
out interference from most other ions.
The glass pH electrode is the nearest
approach to a universal pH indicator
known at present. It works on the
principle of establishing a potential
across a pH sensitive, glass membrane
whose magnitude is proportional to the
difference in pH of the solution
separated by this membrane.
Sawyer, C.N., and McCarty, P.L.
Chem. for San. Eng. 2nd Ed.
McGraw-Hill, NY, 1967
Instruction-Manual
IL 175 PORTO-matic
pH meter Instrumentation
Laboratory, Inc.
Lexington, Mass.
All glass pH indicating electrodes have
a similar basic design. Contained on
one side of an appropriate glass membrane
is a solution of constant pH. In contact
with the other side of this pH sensitive
glass is the solution of unknown pH.
Between the surfaces of the glass mem-
brane, a potential is established which
is proportional to the pH difference of
these solutions. As the pH of one
solution is constant, this developed
potential is a measure of the pH of
the other.
To measure this potential, a half-cell
is Introduced Into both the constant,
internal solution and into the unknown,
external solution. These half-cells are
in turn connected to your pH meter. The
internal reversible half-cell sealed
within the chamber of constant pH Is
almost exclusively a wire of silver-
silver chloride. The external re-
versible half-cell is often silver-
silver chloride. If both the internal
and external electrodes are combined in
a common pH measuring device, the
electrode is a combination pH electrode.
Paoe Ho. 6-14

-------
EFFLUENT MONITORING PROCEDURE: pH Determination of Wastewater and
Wastewater Treatment Plant Effluents
Sections I, VII
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
As the function of these half-cells is to
provide a steady reference voltage
against which voltage changes at the
glass pH sensitive membrane can be re-
ferred, they must be protected from con-
tamination and dilution by the unknown
solutions. This is accomplished by
permanently sealing the internal half-
cell in a separate chamber which makes
electrical contact to the unknown solu-
tion through a porous ceramic plug. This
ceramic plug allows current to flow, but
does not permit exchange of solution to
this chamber. Gradually the KC1 solution
is slowly lost, therefore a filling port
is placed in this electrode so that addi-
tional saturated potassium chloride can
be added.
1. Instrument Calibration
The pH balance control, by adding a
voltage in series with the pH electrode
system, allows the operator to adjust
the meter readout to conform to the pH
of the calibrating buffer. In general,
calibrate the meter in the general range
of the unknown solutions. Appropriate
buffers can be selected (pH 4.0, 6.8,
7.4 and 10.0). Always set the tem-
perature compensator on the instrument
to the temperature of the standard buffer
solution.
For most accurate analysis the pH of the
sample should be determined, and then
buffered solutions of a pH above and be-
low the determined pH should be selected
to re-calibrate the instrument and the
determination of the pH of the sample
repeated for a final reading.
1. The reference chamber of the pH elec-
trode system should always be kept
nearly full of saturated KC1 solution.
Routinely check the level and saturation
VII
Laboratory Analysis
VII
Maintenance Practices
Page No. 6-15

-------
EFFLUENT MONITORING PROCEDURE: pH Determination of Wastewater and
Wastewater Treatment Plant Effluents
TRAINING GUIDE NOTE
of potassium chloride in this reference
chamber and add saturated KC1 if
necessary.
2.	The pH sensitive glass membrane dehy-
drates when removed from water, and
thus it is imperative that dry electrodes
be soaked in buffer or water for several
hours before use. To avoid this break-
in period always keep the glass pH sensi-
tive membrane wet between periods of use.
3.	The buffers are pH standards; do not
contaminate them.
4.	The meter is a battery operated instru-
ment. To conserve the battery life,
turn the instrument off when not in use.
REFERENCES/RESOURCES
Page No. 6-16

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Table 144(1): Preparation of pH Standard Solutions
Standard Solution (Molality)	pH at 25 C'
Primary standards
Potassium hydrogen tartrate
(saturated at 25 C)	3.557
0.05 potassium dihydrogen citrate	3.776
0.05 potassium hydrogen phthalate	4.008
0.025 potassium dihydrogen
phosphate + 0.025 disodium hydrogen
phosphate
0.008695 potassium dihydrogen
phosphate + 0.03043 disodium hydro-
gen phosphate	7.413
0.01 sodium borate decahydrate
(borax)	9.180
0.025 sodium bicarbonate + 0.025
sodium carbonate	10.012
Weight of Chemicals Needed per 1,000
ml of Aqueous Solution at 25 C°
6.4gKHC4H406*
11.41gKH2C6H507
10.12gKHC8H404
6.865 3.388gKH2P04+ + 3.533gNa2HP04+*
1.179gKH2P04+ + 4.302gNa2HP04t#
3.80gNa2B407•10H20#
2.092gNaHC03 + 2.640gNa2C03
Secondary standards
0.05 potassium tetroxalate dlhydrate	1.679
Calcium hydroxide (saturated at 25 C°) 12.454
12.61gKH3C408'2H20
1.5gCa(0H)2*
~Approximate solubility
tDry chemical at 110-130 C° for 2 hr.
^Prepare with freshly boiled and cooled distilled water (carbon dioxide-free)
Page No. 6-17

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NAME
LABORATORY RESULTS
SAMPLE
pH RESULT
Sample 1

Sample 2

Sample 3

Buffer 4

Buffer 9



Page No. 6-19

-------
A PROTOTYPE FOR DEVELOPMENT OF
ROUTINE OPERATIONAL PROCEDURES
for the
COLLECTION AND HANDLING OF BACTERIOLOGICAL
SAMPLES FROM A WASTEWATER TREATMENT FACILITY
as applied 1n
WASTEWATER TREATMENT FACILITIES
and 1n the
MONITORING OF EFFLUENT WASTEWATERS
Developed by the
National Training and Operational Technology Center
Municipal Operations and Training Division
Office of Water Program Operations
U.S. Environmental Protection Agency
W.BA.sa.EMP.la.6.77
Page No. 7-1

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EFFLUENT MONITORING PROCEDURE: Collection and Handling of Bacteriological
Samples
This Procedure was developed by:
NAME	Rocco Russomanno
ADDRESS EPA, OWPO, NTOTC, Cincinnati, Ohio 45268
POSITION Microbiologist-Instructor
EDUCATION AND TECHNICAL BACKGROUND
B.S. University of Massachusetts
5 years Enforcement Action, Rarltan Bay Project, PHS & DI
10 years Instructor, NTOTC, MOTD, OWP, EPA
NAME	Harold L. Jeter
ADDRESS EPA, OWPO, NTOTC, Cincinnati, Ohio 45268
POSITION Chief, Program Support Training Branch
EDUCATION AND TECHNICAL BACKGROUND
A.B. James Milllkln University
M.S. Ohio University
4 years U.S. Army, Clinical Laboratories,
specializing bacteriology
3 years College Instructor - Bacteriology
6 years Research In Sanitary Microbiology
22 years Training of Federal, State and local personnel in
principles and practices of sanitary bacteriology of water
Page No.

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EFFLUENT MONITORING PROCEDURE: Collection and Handling of Bacteriological
Samples
1.	Objective:
Proper technique for the collection and handling of a sample for bacterio-
logical examination taken from a wastewater treatment facility.
2.	Brief Description of Analysis:
After assembly of necessary equipment and travel to the sample site, the
sample is collected in a manner which does not bias the sample. Samples
are collected in a suitable, labeled, sterilized sample bottle which may
contain chemical agents to inactivate chlorine disinfectant or reduce effects
of toxic metals if these are present in the sample.
Bottle handling and filling, using hand or suitable holding device, 1s ac-
complished in a manner to avoid entry of non-representative contamination
which can alter the bacteriological test results.
Cooling or refrigeration, if the sample is held for longer than one hour
before bacteriological laboratory testing, 1s accomplished at less than
10°C but avoiding freezing of the sample. Holding or transporting 1n
this condition can be for no more than 6 hours before the bacteriological
testing must be initiated for which another 2 hours is allowed as pro-
cessing time.
3.	Applicability of this Procedure:
Treatment of Interferences in Samples:
This procedure includes directions for dechlorination of samples sufficient
to act upon samples containing up to 15 mg of residual chlorine per liter.
Also, directions are given for metal detoxification by chelation if these
materials are present or expected in the sample.
Source of Procedure: Standard Methods for the Examination of Water and Waste
water, 14th ed., 1975.

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EFFLUENT MONITORING PROCEDURE: Collection and Handling of Bacteriological
Samples
Laboratory Equipment and Supplies
General Description of Equipment Used 1n the Process
Sample Bottle
Bacteriologlcally Inert; resistant to sterilizing conditions; capacity
at least 100 ml plus air space; containing dechlorlnatlng agent 1f a
sample containing chlorine Is anticipated; and containing a chelating
agent 1f metals are anticipated 1n sample.
Label
Clean and unused; non-smear1ng If wetted; sufficient size for needed
information; can be attached securely to sample bottle.
Marking Device
Non-smear1ng if wetted; permanent marking.
Sampler Device
Unnecessary if bottle can be hand dipped; line, wire, etc.. 1f distance
to sample water Is short; special apparatus 1f distance to sample water
1s sufficient to make line unwieldy or 1f sample water 1s reached with
difficulty as through manholes, ports, etc.
Germicide and Sponge
Disinfecting agent.
Rubber Gloves
Undamaged condition and of proper size.
Container
Ice chest with cover.
Reagents
EDTA (Ethylene d1n1tr1lotetra acetic acid)
Sodium thiosulfate (NagSgOg.SHgO)
Paqe No. 7-5

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EFFLUENT MONITORING PROCEDURE: Collection and Handling of Bacteriological
Samples
Capital Equipment
Autoclave
Providing uniform temperatures up to, and including 12T°C, equipped with
an accurate thermometer, pressure gauges, safety features, saturated steam
power lines and capable of reaching required temperature within 30 minutes.
A1ternately
A pressure cooker can be substituted if:
* effluent pressure gauge
*.thermometer bulb 2.5 cm above water level
Balance
0.1 g sensitivity at load of 160 g.
Oven
Drying and sterilizing. Capable of uniform temperatures and with suitable
thermometer to register accurately In the range 160 - 180 C.
Refrigerator (at laboratory)
Set for 2°-10°C.
Distillation Apparatus, Water
In order of preference, the systems are of stainless steel, quartz, vycor
and pyrex glass. Tin-lined hardware 1s acceptable but because of maintenance
problems 1s best avoided 1n preference to the above. Plumbing should be of
stainless steel, pyrex or plastic PVC material. Storage reservoirs of stain-
less steel and dust protected. Produced water must be of suitable bacterlo-
logic quality (test described 1n Standard Methods, 14th Edition, P. 887.)
Alternately
Distilled water meeting this quality criteria can be purchanged, eliminating
the need for the distillation apparatus.
Washer. Glassware
Operate at 180°C during hot detergent cycle; hot rinse cycle of 6 to 12
successive washings; and final rinse of bacteriologically suitable distilled
or deionlzed water.

-------
ErrLUEHT MONITORING PROCEDURE.: Collection and Handling of Bacteriological Samples
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTE:
A. Presaapling Pro-
cedures
T. Sample bottles
2.	Labels
3.	Label Marker
4.	Sampling Device
1. Check sample bottles for
acceptability
1. Check labels for accepta-
bility
1. Check label marker for
acceptabl1i ty
1. Check sampling service
for condition
la. Suitable material of at least 100 ml capacity.
lb. Bottles should not be chipped, cracked or other-
wise damaged. No deposits or extensive glass
scratches or etched surfaces can be tolerated.
1c. Bottle covers must not be cracked or otherwise
damaged.
la. Must be clean and unused.
lb. Sufficient quantity for number of samples plus
a few extra labels.
1c. Each label must have a means of attachment to
sample bottles. Wire or cord is desirable and
such attachments as scotch tape, electrical tape,
etc. must be avoided as these are affected by
water Immersion.
Id. Labels can vary from that which 1s completely
blank to a type which is required by the facility,
agency, authority, etc.
la. Marker must be of a non-smearing permanent type.
lb. Marker is operable.
la. A number of suitable sampling devices are avail-
able and the function to a) provide weight to
allow the sampling device to reach a depth with-
out drifting; b) provide an anchoring point for
the sterile bottle or chamber; c) have a tripping
mechanism to allow entry of sample to the collec-
tor; and d) provide a means of lowering the device
to depth and returned to surface. Check operation
of each of these areas.
lb. Some types of samplers do not utilize a bottle
but may function with a bulb, bladder, etc. It
will be necessary for the sampler to acquaint
V.A.I.la
(P- 17)
Page No. 7-7

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MONITORI^IGJPROCEDUR^: Collection and Handling of Bacteriological Samples
Page No. 7-8
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
A. Presampling Pro-
cedures (Continued)
5. Germicide
6. Rubber Gloves
7. Chest, Insulated
8. Refrigerator
9. Glassware
1. Availability of germicide
1. Check rubber gloves for
acceptability
1. Check chest for accepta-
bility
1. Check refrigerator for
acceptability
1. Hash all glassware in hot
detergent solution
2. Rinse in hot tap water
himself to the specific device being utilized
at his facility.
la
Provides a means of disinfecting any spillage
of sewage or sample,
lb. Must not be used in a manner where it could find
Its way to contaminate sample, equipment, etc.
Sufficient quantity available (normally about
one pint will suffice for any contingency).
lc
la. Proper sized to fit comfortably.
lb. Must not be punctured.
la. Must be of sufficient size to accoinnodate samples
to be taken.
lb. Must be undamaged so that cold temperature will
be retained inside chest. Must have tight fitting
cover.
lc. Contains ice to quickly chill samples. Must not
have too much water volume since this can compro-
mise sample.
la. Sufficient shelf space for samples. Use of
refrigerator will only be necessary if it is not
possible to run samples immediately upon return
to laboratory.
lb. Temperature setting 2 -10 C.
la. Nontoxic detergent
lb. Be sure all contents and markings are washed
away
2a. At least 6-12 successive rinses to remove
detergent residue
V1.A.5
(p. 18)
V.A.9.1-4
(P. 17)

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EFFLUENT MONITORING PROCEDURE: Collection and Handling of Bacteriological Samples
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
A. Presampling Pro-
cedures (Continued)
3.	Rinse in distilled water
or deionized water
4.	Dry in air
3a. Water must be known to be suitable for bacterio-
logical operations.
3b. 6 to 12 successive rinses recommended to remove
all residues.
4a. No visible spots or scum; glass should be clean
and sparkling.
4b. Glassware must not contain bacteriostatic or
inhibitory residues.
IMPORTANT
The following special conditions may apply to
the sample to be analyzed:
*	If sample is chlorinated effluent which con-
tains copper, zinc, or heavy metals, do opera-
ting procedures A.10, A.ll, and A.12 completely.
*	If sample is unchlorinated effluent which con-
tains copper, zinc, or heavy metals, eliminate
steps: A.10, and A.12.1.
*	If the sample is chlorinated effluent which
does not contain copper, zinc, or heavy metals,
eliminate steps: A.ll and A.12.2.
*	If the sample is unchlorinated and contains no
copper, zinc, or heavy metals, eliminate steps:
A.10, A.ll, A.12.1 and A.12.2.
VI.A.9.3a
(P- 18)
V.A.9.4b
(P- 17)
Paqe No. 7-9

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EFFLUENT MONITORING PROCEDURE: Collection and Handling of Bacteriological Samples
Page No. 7-10
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
A. Presampling Pro-
cedures (Continued)
10. Sodium thiosulfate
solution
11. Ethylene-dinitriTo-
tetra acetic acid
(EDTA) solution
12. Sample bottle
preparation
1.	Weigh 10.0 grams of sodium
thiosulfate
2.	Dissolve 1n 50-60 ml dis-
tilled water
3.	Add distilled water to
bring final volume to
100 ml.
4.	Transfer to labeled bottle
1. Weigh 15.0 grams of EDTA
2.	Dissolve 1n 50-60 ml of
distilled water
3.	Add distilled water to
bring final volume to
100 ml
4.	Transfer to clean labeled
bottle
1. Deliver 0.1 ml or 0.2 ml
thiosulfate solution to
each sample bottle. (.1 ml
to 4 oz. or 120 ml size
and .2 ml to 6-8 oz. or
250 ml size)
la. Used for dechlorination of samples,
lb. Use of trip balance accepted.
2a. 100 ml graduated cylinder satisfactory.
4a. Should be labeled as 10% sodium thiosulfate and
stored in refrigerator.
la. Used for water samples high in copper or zinc,
or wastewater high in heavy metals,
lb. Use of trip balance accepted.
2a. 100 ml graduated cylinder satisfactory.
4a. Labeled as 15% Ethylene dinitrilotetra acetic
acid (EDTA) and stored in refrigerator.
la. Use 1 ml pipet.
lb. Provides adequate sodium thiosulfate for
neutralizing chlorine in sample.
V.A.12.1-6

-------
EFFLUENT MONITORING PROCEDURE: Collection and Handling of Bacteriological Samples
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
A. Presampling Pro-
cedures (Continued)
2.	Deliver .3 ml or .6 ml of
15% EDTA solution to each
sample bottle (.3 ml to
4 oz. or 120 ml. size and
.6 ml. to 6-8 oz. or 250
ml size
3.	Place cover on sample
bottle
4.	Place paper or metal foil
cover over bottle cap or
stopper
5.	Sterilize sample bottles
6.	Store sample bottles in
clean, dry place until
used
2a. Use 1 ml pi pet.
2b. Provides adequate EDTA chelating agent for
metals in sample
2c. Return stock solution of EDTA to refrigerator.
4a. Protects opening of sample bottle from accidental
contamination.
5a. 1 hour at 170°C.

B. Travel: Assembly
Point to Sample Point
1. Initial Sampling
1.	Proceed to initial sample
point
2.	Prepare sample station for
collection of sample
la. Transport equipment with care,
lb. Upon arrival recheck as to correctness of
designated sampling point.
2a. Remove manholes, ports, access panels, etc.,
if necessary.
2b. Note safety hazards at site. It is necessary
to have a partner if potentially hazardous con-
ditions can result in injury or death if another
person is not available for help.

Page No. 7-11

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EFFLUENT MONITORING PROCEDURE: Collection and Handling of Bacteriological Samples
Page No. 7-12
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
C. Sample Collection




1. Spigot or tap
1.
Put on rubber gloves



2.
Flush spigot
2a. Must have direct main connection.
2b. Full flow flush for 2-3 minutes or enough to
clear service line.


3.
Remove hood and cap
from sample bottle
3a. Remove as unit.
3b. Discard slip of paper which is between cap
and bottle.
3c. Protect unit from contamination. Usual method
is to hold cap in left hand (if right-handed)
and have bottle in right hand.


4.
Let sample run into
bottle
4a. No rinsing of bottle. Especially important if
bottle contains sodium thiosulfate or EDTA.
4b. Fill about 3/4 full so that a mixing space is
available for thorough sample mixing prior to
laboratory operations.


5.
Replace cap and hood
on bottle
5a. Secure closure but not excessively tightened
or wedged on bottle.


6.
Label bottle
6a. Fill all items required by local authorities.


7.
Place bottle on ice in
ice chest
7a. Do not immerse bottle in water. Remove ex-
cessive water if present in chest.
7b. Cover chest.

2. River, Stream,
Lake, holding
tank, etc.
1.
Put on rubber gloves.
la. If highly contaminated by direct sewage or
count of water is unknown.

-------
EFFLUENT MONITORING PROCEDURE: Collection and Handling of Bacteriological Samples
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
C. Sample Collection
(Continued)
2.	Remove cap and hood
from sample bottle
3.	Hold bottle near base
4.	Submerge bottle
5.	Replace cap and hood
on bottle
2a. Remove as unit.
2b. Discard slip of paper which is between cap
and bottle.
2c. Protect unit from contamination. Usual method
is to hold cap in left hand (if right-handed)
and have bottle in right hand.
4a. Note current flow of sample site. Bottle filling
operation must be toward flow to avoid contamina-
tion from sampler's hand. If current flow is not
present, the sampler must push the bottle away
from his hand or body to simulate flowing con-
ditions.
4b. Upon entry into water have the neck pointing
downward to prevent surface material from enter-
ing bottle.
4c. When submerged, tilt bottle neck upward to allow
bottle to take in sample water.
4d. If water is shallow the bottle may have to be
held in a hori2ontal position for filling but
the same precautions must be observed to avoid
contamination.
4e. Allow bottle to fill about 3/4 capacity and then
quickly lift out of water. Overfilling or flush-
ing must not occur.
5a. Secure closure but'avoid excessive tightening
or wedging of cap.

Page No. 7-13

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EFFUJEh^MONITORING^^	Collection and Handling of Bacteriological Samples
Page No. 7-14
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
C. Sample Collection
(Continued)
3. Device Collected
Sample
6.	Label bottle
7.	Place bottle on ice in
ice chest
1.	Place sterile sample
bottle in device
2.	Immerse sample device
to depth required
3.	Trip device
4.	Recover device
5.	Remove bottle
6.	Label bottle
7.	Place bottle on ice
in ice chest
6a. Fill all items required by local authorities.
7a. Do not immerse bottle in water. Remove ex-
cessive water if present in chest.
7b. Cover chest.
la. Some devices may employ sterile plastic bags,
containers, etc.
2a. Mark line to indicate depth measurements.
3a. Allow approximately 10 seconds for bottle to
fill or time recommended by manufacturer of
device.
6a. Fill all items required by local authorities.
7a. Do not immerse bottle in water. Remove ex-
cessive water if present in chest.
7b. Cover chest.

D. Sample Handling
i
1. Transport ice chest
to laboratory
la. If sample is bacteridogically processed by
the laboratory within 1 hour of collection,
icing is not required,
lb. If sample is held for over 1 hour, icing is
mandatory.

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EFFLUENT MONITORING PROCEDURE: Collection arid Handling of Bacteriological Samples
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
0. Sample Handling
(Continued)

lc. Sample delivered to laboratory no later than
6 hours from collection time.
Id. Refrigerator can be used for sample holding
or ice chest can be utilized provided that:
a.	Ice is present for holding time
b.	Bottles do not become immersed
in ice water accumulation
c.	Chest remains covered

Page No. 7-15

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EFFLUENT MONITORING PROCEDURE; Collection and Handling of Bacteriological Samples
TRAINING GUIDE
SECTION	TOPIC
I	Introduction
II	Educational Concepts - Mathematics
III	Educational Concepts - Science
IV	Educational Concepts - Communications
V*	Field & Laboratory Equipment
VI*	Field & Laboratory Reagents
VII	Field & Laboratory Analyses
VIII	Safety
IX	Records and Reports
~Training guide materials are presented here under the headings marked*.
These standardized headings are used through this series of procedures.
Page No. 7-16

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AFFLUENT MONITORING PROCEDURE: Collection and Handling of Bacteriological Samples
FIELD & LABORATORY EQUIPMENT
Section v
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
A.1.1a
A.9.1-4
A.9.4b
A.12.1-6
Sample bottles must be composed of a material
which is nontoxic to bacteria, resistant to solvent
action of water, and capable of being sterilized.
Preferably an all glass, ground-glass-stoppered
closure of about 250 ml size is recommended.
Various plastics (polypropylene, Nalgene, etc.)
have been found to meet the specifications above,
and, closures of the screw cap type are acceptable
provided they are, and remain, non toxic to the
sample and provide a tight closure.
Information is applicable to both glass or plastic
sample bottles having the mandatory qualities men-
tioned for sampling use.
Glassware can be checked for bacteriostatic or
inhibitory residues by a bacteriological test
procedure which, like the distilled water suita-
bility test, should be undertaken only by pro-
fessional bacteriologists or 1n laboratories where
this test 1s done on a regular basis.
Wide-mouth, glass-stoppered bottles suggested, but
other styles accepted. Bottle must be heat stable
to sterilizing conditions and not be toxic or
nutritive to organisms natural to the sample.
If glass-stoppered bottles are used, a strip of
paper should be placed in the neck of the bottle
before placing the stopper 1n place in preparation
for sterilization. This prevents the glass stopper
from "freezing" in place during sterilization. The
paper strip 1s discarded at the time of sample
collection.
Standard Methods for the
Examination of Water and
Wastewater, 14th ed.,
1976, APHA, New York,
New York, p. 885.
Standard Methods for the
Examination of Water and
Wastewater, 14th ed.,
1976, APHA, New York,
New York, p. 884, p. 904.
Page No. 7-17

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EFFLUENT MONITORING PROCEDURE: Collection and Handling of Bacteriological Samples
FIELD & LABORATORY REAGENTS
Section VI
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
A.5
A.9.3a
A large number of preparations can be used as
germicides varying from dilutions made from pur-
chased concentrates or laboratory preparations
made by the worker. A few of the more easily made
germicides are as follows:
1. Ethyl Alcohol - 70% solution
Examples:
7 ml
70 ml
700 ml
ethanol + 3 ml distilled water
ethanol + 30 ml distilled water
ethanol + 300 ml distilled water
2.	Isopropyl Alcohol - 70% solution
3.	Ethyl Alcohol (70% solution) containing
1% Iodine crystals
Examples:
7 ml ethanol + 3 ml distilled water;
0.1 gram Iodine crystals added to the
ethanol-water solution
70 ml ethanol + 30 ml distilled water
1.0 grams of Iodine crystals added to
the ethanol-water solution
4.	Isopropyl Alcohol (70% solution) containing
1% Iodine crystals
5.	Aqueous KI (potassium iodide) 1% contain-
ing 1% Iodine
Examples:
1 gram KI + 100 ml distilled water,
mix until dissolved, then add 1 gram
of Iodine crystals and mix until dis-
solved.
10 grams KI + 1000 ml (or 1 liter)
of distilled water, mix until dis-
solved, then add 10 grams of Iodine
crystals and mix until dissolved.
Distilled water must not contain substances pre-
venting bacterial growth or be highly nutritive.
There are required procedures to testing distilled
water and should be undertaken only by professional!
bacteriologists or in laboratories where this Is
done regularly.
Standard Methods for the
Examination of Water and
Wastewater, 14th ed.»
1976, APHA, New York,
New York, p. 881-891.
Page No. 7-18

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A PROTOTYPE FOR DEVELOPMENT OF
ROUTINE OPERATIONAL PROCEDURES
for the
FECAL COLI FORM TEST
by the
MULTIPLE DILUTION TUBE METHOD
as applied in
WASTEWATER TREATMENT FACILITIES
arid in the
MONITORING OF EFFLUENT WASTEWATERS
Developed by the
National Training and Operational Technology Center
Municipal Operations and Training Division
Office of Water Program Operations
U.S. Environmental Protection Agency
W.8A.EMP.Jc.6.77
Page No. 8-1

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EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Multiple
Dilution Tube Method
This Procedure was developed by:
NAME Harold L. Jeter
ADDRESS EPA, OWPO, NTOTC, Cincinnati, Ohio 45268
POSITION Chief, Program Support Training Branch
EDUCATION & TECHNICAL BACKGROUND
AB - James Ml11ikin University
MA - Ohio University
4	years - US Army; Clinical Laboratory; specializing
bacteriology
3 years College Instructor - Bacteriology
6 years Research 1n Sanitary Microbiology
21 years Training of Federal, State and Local personnel
in principles and practice of sanitary bacteriology water
NAME Rocco Russomanno
ADDRESS EPA, OWPO, NTOTC, Cincinnati, Ohio 45268
POSITION Microbiologist-Instructor
EDUCATION & TECHNICAL BACKGROUND
BS - University of Massachusetts
5	years Enforcement Action, Rarltan Bay Project, PHS & DI
9 years Microbiologist-Instructor
Page No. 8-3

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FECAL COLIFORM TUST
Multiple Dilution Tube (MPN) Method
Flow Sheet
SAMPLE
LL.STU
5 tubes of each of a decimal at>ries of
3 or more sample volumes. (Kach
successive volume is 1/10 the previous;
volume.)
Incubate 24 + 2 hrs. at 3b°C + 0. 5°C
Gas (+)
Gas (-)
ECB
Incubate in water bath 24 hrs. + 2 hrs.
at 44. 5°C + 0. 2°C
Incubate 24 hrs. + 2 hrs. at 35°C + 0. 5°C
Gas (+)
Fecal Collforms
Present
No Gas (-)
Fecal Collforms
Not Present
Gas (+)
No Gas (-)
Fecal Coltforms
Not Present
ECB
Incubate In water bath 24 hrs. + 2 hrs. at 44. 5 +0. 2°C
Gas (+)
Fecal Collforms
Present
J
Codify results based on No. Gas + tubes in EC IJroth
I
Determine MPN Index
I
Apply correction factor as required
Record results as Fecal Colifonnc/100 ml
Gas (-)
Fecal Coliforms
Not Present
Pane No. 8-4
Report results as prescribed under
¦FOBS or other regulatory requirements

-------
EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Multiple
Dilution Tube Method
1.	In wastewater effluent quality control, the objective of the test may be one
or both of the following:
a.	To determine whether the bacteriological quality of the effluent meets
quality requirements set by law or by regulatory authority.
b.	To determine overall efficiency of the treatment process in reducing the
bacterial content of the wastewater being treated.
2.	Brief Description of Analysis:
Three or more decimal series dilutions of a sample (for example: five fermenta-
tion tubes with 10-ml portions, another five tubes with 1-ml portions, etc.)
are inoculated into lactose lauryl sulfate trytose broth (LST) and incubated
at 35°C + 0.5OC. After 24 hours and again after 48 hours the LST cultures
are examined and results recorded for gas production. Cultures showing gas
are transferred at each examination time to EC Broth (EC) fermentation tubes
and incubated at 44.50 + 0.2OC in a water bath. EC cultures are examined for
evidence of gas production after 24 hours. At the end of the overall incubation
period, results are summarized as positive or negative and coded to represent
the number of EC gas-positive tubes for each series. A Table of Most Probable
Numbers (MPN) is used with the coded results to determine an MPN Index. This
index is corrected (if necessary, since the table is for 5-tube, 10, 1.0, and
0.1 ml series only) to agree with the actual sample volumes indicated. The
final result is recorded as Fecal Coliforms per 100 ml of sample.
3.	Applicability of this Procedure:
Range of Concentration:
If these dilutions are used
These ranges
are covered
1.0; 0.1; 0.01;0.001
20 to
160,000
0.1; 0.01; 0.001; 0.0001
200 to
1,600,000
0.01; 0.001; 0.0001; 0.00001
2,000 to
16,000,000
0.001; 0.0001; 0.00001; 0.000001
20,000 to
160,000,000
b. Pretreatment of Samples
In accordance with Standard Methods, 14th Ed. (p. 904), and as outlined in
EMP, "Collection and Handling of Bacteriological Samples".
This procedure conforms to the fecal coliform test as described 1n
Standard Methods for the Examination of Mater and Wastewater, 14th
Ed. (1975), p. 922 ff.
Page No. 8-5

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EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Multiple
Dilution Tube Method
General description of equipment and supplies used in the test analysis
Capital Equipment
Autoclave, providing uniform temperatures up to and including 121°C,
equipped with an accurate thermometer, pressure gauges, saturated
steam power lines and capable of reaching required temperature within
30 minutes.
Balance, 0.1 g sensitivity at load of 150 g
Air incubator to operate at 35°C + 0.5°C
Incubator, waterbath, to operate at 44.5° + 0.2°C and to accommodate
tube racks as described separately
Oven, *hot-air sterilizing, to give uniform temperatures and with suitable
thermometer to register accurately in range of 160~180°C
pH meter, accurate to at least 0.1 pH unit, with standard pH reference
solution(s)
Water distillation apparatus (glass or block tin), or source of distilled water
suitable for bacteriological operations.
Reusable Supplies
Apron or coat suitable for laboratory
Baskets, wire for discarded cultures
Bottles, dilution*, 6-oz. screw caps, with 99ml volume level etched on
one side
Bottles, sample*, preferred characteristics being 250 ml (6-8 oz.),
wide mouth, glass stopper
Bottle, squeeze type, with disinfecting solution
Burner, gas, Bunsen burner type
Cans, plpet, aluminum or steel; not copper (If plastic, or other type
of prepackaged disposable pipets are used, this Item 1s unnecessary.)
Metal caps* to fit 20 x 150 irm culture tubes
Pan, to receive discarded contaminated pipets and glassware (must contain
disinfectant before use)
Inoculation loop, 3 mm diameter loop of nlchrome or plat1num-1r1dium
wire, 26 B&S gauge, in holder
Pipets*, 1 ml, with 0.1 ml graduations, Mohr type preferred, sterile,
cotton plugged, glass or disposable plastic
Paae No. 8-6

-------
Racks, culture type* 10 x 5 openings, to accept tubes at least 20 mm
in diameter
Sponge, for cleaning desk top
Tubes, culture*, 150 x 18 mm
Tubes, fermentation*, 75 x 10 mm vials to be inverted in culture tubes
Supplies Used Up in the Analysis (must be replaced when stocks get low)
Distilled water, suitable for bacteriological cultures (note distillation
apparatus required in capital equipment)
EC Broth, Dehydrated (recommend purchase of 1/4-lb. units)
Lactose Lauryl Sulfate Tryptose Broth, Dehydrated (recommend purchase of
1-lb. units)
Potassium Dihydrogen Phosphate (K^PO^) (recommend purchase of 1/4 lb. units)
Disinfectant, for bench tops. (Use household bleach solution prepared
according to instructions on bottle)
Wax pencils (recommend soft wax equivalent to Blaisdell 169T)
EDTA (ethylene clinitrilotetraacetic acid)
Sodium thiosulfate (Na^O^ f^O)
*Iterns marked are needed in quantities or require size or space allowances
which cannot be specified here, as they vary according to the daily analysis
schedule. As a rule-of-thumb, space/size or quantity requirements should be
at least 3 times the normal daily requirements. For further information on
specifications for equipment and supplies, see the Microbiology Section of
the current edition of "Standard Methods for the Examination of Water and
Wastewater."
Page No. 8-7

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EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Multiple
¦	Dilution Tube (MPN) Method
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
A. Pre-Test Procedures
1. 35#C Incubator set-up,
adjustment
1. Place 35°C incubator in
permanent location.
2. Install thermometer.
3. Install shallow pan of
water 1n bottom of incu-
bator.
4.	Connect incubator to
electric power source.
5.	Adjust temperature until
stabilized at required
temperature.
6. Operate bacteriological
incubator continuously.
Aa. All pretest procedures completed before starting
other first-day procedures.
la. Out of drafts or places where it will be in direct
sunlight part of day.
lb. Location convenient to laboratory bench.
1c. Convenient source of electric power.
2a.
2b.
2c.
3a.
Thermometer functions at least in 30°-40°C range
and have intervals of 0.5° or less indicated.
Meets NBS standards.
Location should be central in incubator.
Mercury bulb thermometer should be fitted with
cork or rubber stopper and mounted in small bottle
filled with liquid (glycerine, water, or mineral
oil).
In most laboratory incubators a pan having about
1 square foot of area, with water about 1 inch
deep, is satisfactory.
3b. Maintains condition of saturated relative humidity,
required in bacteriological incubator.
3c. Requires dally check, with addition of water as
necessary, to keep water in pan at all times.
4a. Many incubators have pilot light to indicate
power turned on.
5a. Manufacturer's instructions for method of
temperature adjustment.
5b. Operation must be at 35 + 0.5°C.
5c. Should allow about 1 hour between adjustments.
6a. Requires daily check with written temperature
record, with adjustment and water addition as
necessary.
V.A.I.1
(P. 41)
V.A.I.2
(P. 41)
V.A.I.3
(P. 41)
V.A.I.5
(P- 41)
V.A.I.6
(P- 41)
Page No. 8-9

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EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Multiple
Dilution Tube (MPN) Method
Page No. 8-10
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
2. Water bath incubator
setup* adjustment
1.	Place water bath incubator
in permanent location.
2.	Put water in water bath.
3.	Install thermometer.
4.	Connect water bath incu-
bator to electric power
source and turn on.
5.	Adjust temperature until
stabilized at required
temperature.
6.	Operate water bath incu-
bator continuously.
la. On bench or table surface.
lb. Out of drafts or place in which it will be in
direct sunlight part of day.
lc. Location convenient to laboratory bench.
Id. Convenient source of electric power.
2a. Distilled or deionized water preferred, tap water
accepted.
2b. Should be 2-21/2 inches deep above the platform on
which the racks of cultures will be placed. Water
must be deep enough that when racks of cultures
are placed in the water bath the water is as high
on the tubes as the top as the culture medium in-
side the tubes. Yet it must not be so deep as to
let the tubes float out of the racks or reach the
cap.
3a. Functions at least in 40°-50°C range. Meets NBS
standards. Have at least 0.1'C increment markings.
3b. Host water baths provide for corner location for
thermometer (for protection from breakage).
4a. Pilot light should come on.
5a. Manufacturer's instructions for location and
method of temperature adjustment.
5b. Operation must be at 44.5 + 0.2°C.
5c. Allow about 1 hour between adjustments.
6a. Requires daily check with written temperature
record, with adjustment as necessary.
6b. Requires daily check of water level and addition
of more as needed.
6c. With tap water in water bath, may require
periodic scum removal from inner walls.

-------
EFFLUENT MONITORING PROCEDURE: pecal Coliform Test by the Multiple
™¦—>—¦	Dilution Tube (MPN) Method
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPEC IFICATIONS
TRAINING
GUIDE NOTES
3. Oven, sterilizer,
setup
4. Autoclave setup
1.	Place oven sterilizer in
permanent location.
2.	Install thermometer.
3.	Connect oven sterilizer to
power source and turn on.
4.	Adjust temperature to
stabilize at required
temperature.
5.	Operate oven sterilizer
only when needed. Turn
off when not in use.
Install and operate auto-
clave according to manu-
facturer's instructions.
la. Convenient to source of electric power; usually
on table or bench.
2a. Should indicate the 160°-180°C range, be accurate
within this interval, and be marked in 1.0 degree
intervals.
3a. Usually has pilot light to indicate power on.
4a. Operated as near to 170 C as possible; not lower
than 160 nor higher than 180 C .
5a.
5b.
5c.
5d.
la.
lb.
lc.
Id.
le.
If.
Turned ON in advance of need to permit reaching
required temperature before introducing material
to be sterilized.
Oven sterilizer used to sterilize dry glassware,
metal objects.
Oven sterilizer not used with culture media,
solution, plastics, rubber objects, or with
anything containing or including these.
Paper-wrapped glass pi pets may be sterilized in
oven sterilizer.
Autoclaves extremely variable in design and
operation; also, potentially dangerous.
Used to sterilize objects made of, or including
liquids, rubber, culture media.
Glassware may be autoclave sterilized but must be
dried afterward.
Most plastics not sterilized in autoclave;
plastics usually require chemical sterilizers.
Autoclave usually operated at 121 C for 15 min.
Sterilized media must be removed from autoclave
as soon as possible after autoclave is reopened.
V.A.3.1-5
(p. 41)
V.A.4.1
(p. 41)
Page No. 8-11

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EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Multiple
Dilution Tube (HPN) Method
Page No. 8-12
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
5. Water distillation
equipment
6. pH meter
7. Glassware
1.	Install and operate in
accordance with manu-
facturer's instructions.
2.	Operate continuously or
intermittently as required
to maintain adequate
supplies of distilled
water.
1. Have unit available and
operate in accordance with
procedures described 1n
other lab procedures.
1.	Wash all glassware in hot
detergent solution;
2.	Rinse at least once in
hot tap water;
3.	Rinse 1n distilled water,
at least 6 successive
times, and,
la. Must produce distilled water meeting quality
requirements for bacteriological tests.
2a.
2b.
4. Dry in air.
Reserve supplies kept in borosilicate glass
carboys or in plastic carboys made of material
which will not dissolve substances which will
affect growth of bacteria.
Same distillation apparatus used for bacterio-
logical purposes may be used for chemical
reagents.
la. Unit for pH check on finished culture media,
lb. Used in preparation of stock solution of
potassium dihydrogen phosphate.
la. Nontoxic detergent
lb. Be sure all_ contents and markings are washed away.
4a. No visible spots or scum; glass should be clean,
and sparkling.
4b. Glassware suitable for use in bacteriological
operations.
V.A.5.1-2
(p. 42)
V.A.6.1
(P. 42)
V.A.7.1-4a
(p. 42)
V.A.7.4b
(P- 42)

-------
EFKJfeNT MONITORING PROCEDURE: Fecal Coliform Test by the Multiple
Dilution Tube (MPN) Method
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
8.	Sodium thiosulfate
solution
9.	Etylenedlnltrllotetra-
acetic acid (EDTA)
solution
1.	Weigh 10.0 grams of sodium
thiosulfate.
2.	Dissolve in 50-60 ml dis-
tilled water.
3.	Add distilled water to
bring final volume to
100 ml.
4.	Transfer to labeled bottle.
1. Weigh 15.0 grams of EDTA.
The following special conditions may apply to the
sample to be analyzed:
If the sample is chlorinated effluent which
contains copper, zinc, or heavy metals, do
operating procedures A.8, A.9 and A.10 completely
If the sample is unchlorinated effluent which
contains copper, zinc, or heavy metals, elimi-
nate steps: A.8 and A.10.1.
If the sample is chlorinated effluent which
does not contain copper, zinc, or heavy metals,
eliminate steps: A.9 and A.10.2.
If the sample is unchlorinated and contains no
copper, zinc, or heavy metals, eliminate steps:
A.8, A.9, A.10.1 and A.10.2.
la. Used for dechlorination of samples,
lb. Use of trip balance accepted.
2a. 100 ml graduated cylinder satisfactory.
4a. Labeled as 102 sodium thiosulfate and stored in
refrigerator.
la. Used for water samples high in copper or zinc or
wastewater samples high in heavy metals,
lb. Use of trip balance accepted.

Page No. 8-13

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EFFLUENT MONITORING PROCEDURF: Fecal Coliform Test by the Multiple
Tube (MPN) Method
Page No. 8-14
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
10. Sample bottle
preparation
2.	Dissolve in 50-60 ml dis-
tilled water.
3.	Add distilled water to
bring final volume to
100 ml.
4.	Transfer to labeled clean
bottle.
1.	Deliver 0.1 ml or .2 ml
of 10* sodium thlosulfate
solution to each sample
bottle. (.1 ml to 4 ounce
or 120 ml size and .2 ml
to 6-8 ounce or 250 ml
size)
2.	Deliver .3 ml or .6 ml of
151 EDTA solution to each
sample bottle (.3 ml to
4 ounce or 120 ml size and
.6 ml to 6-8 ounce or
250 ml size).
3.	Place cover on sample
bottle.
4.	Place paper or metal foil
cover over bottle cap or
stopper.
5.	Sterilize sample bottles 1n
sterilizing oven.
2a. A 100 ml graduated cylinder is satisfactory.
4a, The bottle should be labeled as 15% Ethylene-
dinitrilotetraacetic acid (EDTA) and stored
in refrigerator.
la. Use 1 ml plpet.
lb. Provides adequate sodium thlosulfate for
neutralizing chlorine in sample.
lc. Return stock sodium thlosulfate solution to
refrigerator.
2a. Use 1 ml plpet.
2b. Provides adequate EDTA chelating agent for metals
in sample.
2c. Return stock solution of EDTA to refrigerator.
4a. Protects opening of sample bottle from accidental
contamination.
5a. One hour at 170°C. (See A.3)
V.A.10.1-6
(p. 42)

-------
EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Multiple
Dilution Tube (MPN) Method
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
11. Pi pet preparation
6. Store sample bottles in
clean, dry place until
used.
1. Inspect plpets to be pre-
pared for use; discard and
destroy all having chipped
or cracked tips.
2.	Insert plug of non-
absorbent cotton into
mouthpiece of each clean,
dry pipet.
3.	Place a layer of glass wool
or several layers of paper
padding In bottom of plpet
can.
4.	Place 18-24 pipets in each
pipet can, delivery tip
down.
5.	Sterilize cans of pi pets-
pi pets in oven.
6.	Store cans in clean, dry
place until used.
7.	When can of pipets is
opened for first use, pass
the exposed ends of the
pipets through flame,
slowly.
la. Cleanliness of pipet must be equivalent to
glassware.
lb. For protection of user when pipetting sample.
2a. Cotton plug must be tight enough to prevent easy
removal, either by the pipetting action or by
handling, and yet loose enough to permit easy
air movement through the plug.
3a. For protection of pipet delivery tips.
4a. Permits removal of sterile pipets from can
without contamination by user.
5a. 1 hour at 170°C. (See A.3 of procedures)
6a. Laboratory cabinet or drawer recommended.
7a. Burns off excess cotton sticking out of pipet
mouthpiece.
7b. Cover kept on can at all times except when
samples are being inoculated.
V.A.ll.1-6
(p. 42)
V.A.ll.7
(P. 43)
Page No. 8-15

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EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Multiple
———————"—Dilution Tube (MPN) Method
Page No. 8-16
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
12. Dilution water blanks
1.	Prepare stock solution of
potassium dihydrogen phos-
phate (KH2P04}; dissolve
34.0 grams of the KH2P04
in 500 ml distilled water.
Adjust to pH 7.2 with
IN NaOH, and dilute to 1
liter with distilled water
2.	Prepare stock solution of
magnesium sulfate (MgSO^.
7^0) by dissolving 50
grains of this chemical In
500-600 mis of distilled
water and, after complete
dissolving, bring the
final volume to 1 liter in
a volumetric flask.
3.	Prepare working solution
of dilution water by add-
ing 1.25 ml KH2P04
and 5 ml of the magnesium
sulfate stock solution to
each liter of distilled
water to be made up as
dilution water.
la. Distilled water may be measured in 500 ml
graduated cylinder,
lb. Finished solution labeled "Stock KH?P0. for
Dilution Water."
lc. Stored in refrigerator.
Id. Discard stock solution and prepare new solution
if mold appears.
3a. 5 ml pi pet satisfactory for 1 liter amounts of
dilution water. 10 ml pipet better when several
liters are being made.
3b. 1-liter graduated cylinder satisfactory for
measurement of distilled water.
3c. Use seperate pipets for each solution to prevent
contamination.
V.A.12.1.Id
(p. 43)

-------
EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Multiple
Dilution Tube (MPN) Method
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
13. Preparation of Lactose
Lauryl Sulfate Tryp-
tose Fermentation
Broth (LLSTB)
3.	Deliver enough working
solution to each dilution
water bottle so that after
sterilization the bottles
will contain 99 + 2 ml of
dilution water.
4.	Place caps on dilution
bottles loosely.
5.	Sterilize in autoclave.
6.	Promptly remove from auto-
clave, tighten bottle caps
cool to room temperature.
7.	Store in cool place.
Weigh 35.6 grams of dehy-
drated Lactose Lauryl
Sulfate Tryptose Broth.
Close cover of bottle of
dehydrated medium tightly
after removal.
2.	Dissolve in 1 liter dis-
tilled water»
3.	Place 10 ml of the solu-
tion of prepared LLSTB in
each culture tube.
3a. 100 ml graduated cylinder ordinarily satisfactory
Pipetting machine desirable but not mandatory.
3b. Amount cannot be stated exactly, as sterilization
evaporation differs from one autoclave to another,
Conmonly, about 102 ml is required.
5a. 15 minutes at 1210C.
steam evacuation.
Use "slow-vent" mode of
7a. Dilution water ready for use. May be stored
indefinitely in screw-capped bottles.
la. Dehydrated media take moisture out of air;
can become caked,
lb. Caked media unsatisfactory; should be discarded.
1c. Prepares 100 tubes (enough for 5 tests based on
4 rows of 5 tubes each).
2a. Gentle heat (no boiling) if necessary to com-
plete dissolving medium.
3a. Use 150 x 18 mm tubes.
3b. 10 ml pipet, automatic pipetter, or funnel hose
and pinchcock assembly are acceptable.
3c. Accuracy of delivery: +0.5 ml.
V.A.12.3
(P- 43)
V.A.12.4
(p. 43)
V.A.12.5
(p. 43)
V.A.12.7
(P- 43)
V.A.13.3b
(P. 43)
Page No. 8-17

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EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Multiple
Dilution Tube (MPN) Method
Page No. 8-18
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
Preparation of Lactose
Lauryl Sulfate Tryptose
Fermentation Broth
(LLSTB) (Continued)
,
4.	Insert one fermentation
vial Into each tube of
medium, open end down.
5.	Place tube cap on each
tube culture medium.
6.	Sterilize 1n autoclave.
7.	Cool medium to room tem-
perature.
8.	Check pH of finished
medium.
9.	If final pH not satis-
factory. discard medium
and prepare new batch with
pH adjustment before
sterilization.
10. Store medium in cool dark
place.
4a. Tubes and vials previously washed as indicated
A.7.1-4.
4b. Use 75 x 10 mm tubes.
5a. After all tubes have been filled and have
individual vial.
6a. Within 1 hour after medium prepared.
6b. Sterilization at 121 C for 15 minutes.
6c. Medium must be removed from autoclave as soon
as possible after pressure has returned to
normal. Use "slow-vent" mode of steam removal.
7a. Medium ready for use when cool and individual
vials are completely filled with fluid. No
bubbles must be present.
8a. Should be pH 6.8 - 7.0.
9a. pH value ordinarily drops about 0.2 pH unit.
10a. Not in refrigerator. Usually in laboratory
cabinet in darkness.
10b. May be stored up to 1 week if evaporation not
more than 10%.

-------
FFFLUENT MONITORING PROCEDURE: ^cal Coliform Test by the Multiple
—^™———————1— Dilution Tube (MPN) Method
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
14. Preparation of EC
Broth
1.	Weigh 37.0 grams of dehy-
drated EC Broth. Close
cover of bottle of
dehydrated medium tightly
after removal.
2.	Dissolve in 1 liter dis-
tilled water.
3.	Place 10 ml of the solu-
tion of prepared EC Broth
in each culture tube.
4.	Insert one fermentation
vial into each tube of
medium, open end down.
5.	Place tube cap on each
tube of culture medium.
6.	Sterilize in autoclave.
7.	Cool medium to room
temperature.
8.	Check pH of finished
medium.
9.	If out of range 6.8 - 7.0
discard and prepare again
with prior adjustment of
pH with IN NaOH or HC1.
10. Store medium in cool dark
place.
la. Dehydrated media take moisture out of air, be-
come caked.
lb. Caked media unsatisfactory; discard.
1c. Prepares 100 tubes; this is enough for four to
five tests.
2a. Gentle heat if necessary. No boiling.
3a. Use 150 x 18 mm tubes.
3b. 10 ml pipet, automatic pipetter, or funnel hose
and pinchcock assembly are acceptable.
3c. Accuracy of deliver +0.5 ml
4a. Tubes and vials previously washed as indicated
in A.7.1-4.
4b. Use 75 x 10 mm tubes.
5a. After all tubes have been filled and vials in-
serted.
6a. After all tubes have been capped.
6b. Sterilization at 121 C for 15 minutes.
6c. Medium must be removed from autoclave as soon
as possible after pressure has returned to normal.
7a. Medium ready for use when cool.and individual
vials are completely filled with fluid. No
bubbles must be present.
8a. Should be pH 6.9.
9a. Before sterilization most media should be ad-
justed to 0.2 pH units higher than pH value
expected of the sterile medium.
10a. Not in refrigerator. Usually in laboratory
cabinet in darkness.

10b. May be stored up to 1 week if evaporation not
more than 10%;
Page No. 8-19

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EFFLUENT MONITORING PROCEDURF: Fecal Coliform Test by the Multiple
" Dilution Tube (MPN) Method
Page No. 8-20
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
15. Final equipment and
supply check
1.	Check to be sure that all
equipment and supplies,
solutions, and prepared
media are ready before
starting sample examina-
tion.
2.	Make preparations or ad-
justments as necessary
before starting test.
la. Check general list of equipment and supplies,
lb. Each test requires (with 4 sample volumes per
test) 20 tubes LLSTB;
10-15 tubes EC Broth
1 sample bottle;
1-5 1-ml pipets, sterile; and
1-3 99-ml bottles sterile dilution water

B. First-day Procedures
1.	Equipment Maintenance
2.	Sample collection
1.	Check, record, and adjust
incubator temperature.
2.	Add water to pan in incu-
bator as necessary
1.	Collect sample.
2.	Record sampling informa-
tion
3.	Transport sample to
laboratory
la. See A.1.1-6.
la. Locations as selected by plant management.
lb. Sampling methods as described in procedure
"Sample Collection and Handling for Bacteriologi-
cal Tests" or in Standard Methods.
2a. Most plants have sample tag of some type which
includes such information as date, time, place
of sampling, name of sample collector, and other
information as may be required.
3a. Taken to laboratory without delay.
3b. Samples iced if delay of starting sample test
is greater than one hour. No more than 6 hours
of transportation time is allowed.

-------
EFFUJENTHONITORING^P^	Fecal Coliform Test by the Multiple
Dilution Tube (MPN) Method
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
B. First-day Procedures
(Continued)
3. Preparation of
laboratory data sheet
1.	Fill in data sheet to
show sample information.
2.	Select sample inoculation
volumes.
3.	Enter information in
laboratory data sheet to
show sample inoculation
volume for each series
(row) of 5 tubes.
la. Needed information should be on sample collection
tag.
lb. Most data sheets show at least source, date,
time of collection, name of sampler, name of
analyst, laboratory sample number assigned.
2a. According to fecal coliform density range pre-
dicted for the sample.
2b. For fecal coliforms per 100 ml in the range
from to inoculate 5 tubes each of ml
20- 160,000 1.0, 0.1, 0.01 0.001
200- 1,600,000 0.1 0.01, 0.001 0.0001
2,000- 16,000,000 .01 .001, .0001 .00001
20,000- 160,000,000 .001 .0001, .00001, .000001
2c. For chlorinated effluents, 1.0, 0.1, 0.01, and
0.001 ml sample portions are recommended.
2d. For raw (untreated) sewage, use sample portions
of 0.0001, 0.00001, 0.000001, and 0.0000001 ml.
2e. For other waters, other combinations of sample
volumes may be required, particularly in en-
vironmental waters receiving raw or incompletely
treated sewage. It may be necessary to conduct
exploratory tests.
3a. Recommend showing sample inoculation volumes in
ml or decimal amounts.
VII.B.3.1
(P- 44)
VII.B.3.2
(p. 44)
Page No. 8-21

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EFFLUENT MONITORING PROCEDURE:
Fecal Coliform Test by the Multiple
Dilution Tube (MPN) Method
Page No. 8-22
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
B. First-day Procedures



(Continued)



4. Lab bench
1. Disinfect laboratory
la. Sponge and disinfectant; paper toweling.

disinfection
bench; wipe dry.


5. Assembly and label-
1. Place 5 tubes of Lactose
la. If more than one sample is being tested, rack

ing of culture medium
Lauryl Sulfate Tryptose
with 5 x 10 openings can be used to set up two

Broth (LLSTB) in each of
tests.


4 rows in culture tube



rack. (20 total tubes)



2. Label tubes of culture
2a. Use labeling code.
VII.B.5.2

medium to show sample
2b. Label every tube. Only the experienced worker
(P. 45)

number, sample volume,
should take short-cuts in labeling.

and position of tube in
2c. Use wax pencil. Soft wax equivalent to


the series of 5 tubes per
Blaisdell 169T is suggested.


sample volume.

6. Sample inoculation
1. Shake sample vigorously.
la. At least 25 shakes over space of at least 1 foot
I.B.6.1.1
(with dilution as

in 10 seconds or less.
(p. 34)
required)



2. Deliver Into the labeled
2a. Use sterile 1 ml pi pets.
VII.B.6.2-3

LLSTB tubes the sample

(p. 45)

portions previously


selected.

-------
EF \ENT MONITORING PROCEDURE: Fecal Conform Test by > Multiple
—' 	__ Dilution Tube (MPN) NeuJu
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
B. First-day Procedures
(Continued)
3. Each tine a sample di-
lution Is prepared, shake
vigorously, as with the
original sample.
2b. Table of sample portions
To get
i«ll
1.0
0.1
0.01
0.001
0.0001
0.00001
0.000001
Deliver
10)
100)
1000)
10000)
100000)
1000000)
ML
1.0
0.1
1.0
0.1
1.0
0.1
1.0
From
fsawple preparations)
original sample
original sample
1:100 dilution
1:100 dilution
1:10000 dilution
1:10000 dilution
1:1000000 dilution
2c. Dilutions of original samples
To get
1:100
1:10000
1:1000000
Deliver to
99-nl blank
1 ml
1 al
1 ml
From
original sample
1:100 dilution
1:10000 dilution
3a. At least 25 shakes over space of at least 1 foot
In 10 seconds or less.
Pnnn No. fl-?3

-------
FLUENT MONITORING PROCEDURE:
Fecal Coliform Test by the Multiple
Dilution Tube (MPN) Method
Page No. 8-24
-IRATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
Incubation
Processing used
glassware
Lab bench
disinfection
1.	After completion of sample
inoculation into LLSTB,
shake rack of cultures
qentlv.
2.	Place rack(s) of cultures -1n
Incubator.
1.	Drain Sample bottles,
dilution bottles, and plpets
Into sink.
2.	Wash and dry bottles, plpets
1. Disinfect laboratory bench
top; wipe dry.
la. Mixes sample with culture medium.
lb. Avoid shaking air into fermentation vials.
2a. 24 hours + 2 hours at 35 + 0.5°C
la. Sterilization unnecessary.
2a. Meets original cleanliness requirements of
glassware.
2b. Glassware ready for reuse.
la. Sponge, disinfectant, paper toweling.

24-hour Procedures
Equipment
maintenance
Disinfection
Reading and record-
ing of results.
1.	Check, record, and adjust
incubator temperature.
2.	Add water to pan in incu-
bator as necessary.
1. Disinfect laboratory bench
top; wipe dry.
1.	Remove rack(s) of culture(s)
from incubator to lab bench.
2.	Shake culture rack gently.
la. See A.l.1-6
la. See B.4.1
2a. Hastens release of gas in supersaturated cultures.
2b. )st not shake air Into fermentation vials.

-------
EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Multiple
—' Dilution Tube (MPN) Method
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
4. Transfers
3. Examine each tube for gas
production and record re-
sults on data sheet.
1.	Label and assemble tubes
of EC Broth.
2.	Transfer each gas-positive
tube of LLSTB to a labeled
tube of EC Broth.
3.	Place each inoculated tube
of EC Broth in a separate
rack, in same relative
position as original gas-
positive LLSTB tubes in rack.
4.	After each transfer, place
original positive LLSTB
tube in discard basket.
3a. If present, gas will be trapped in the fermenta-
tion vial.
3b. Gas in any quantity is a positive test.
3c. Vials witn no gas are a negative test.
3d. Each result appears on line corresponding with
the tube label.
3e. All results appear under the "24" of the LLSTB
column.
3f. Plus sign (+) means a gas-positive tube.
3g. Minus sign (-) means a gas-negative tube.
la. One tube for each LLSTB gas-positive tube.
lb. Each EC Broth tube label corresponds with label
on gas-positive LLSTB tube.
lc. Labeled EC Broth tubes assembled in a culture
tube rack in same relative position as gas-
positive LLSTB tubes in their rack.
2a. Label on inoculated tube of EC Broth is the same
as the label on the tube of LLSTB from which the
transfer is made.
2b. 3-mm inoculation loop.
2c. Loop flame-sterilized before use and between
successive transfers.
2d. One loopful per transfer.
III.C.3.3
(p. 39)
VII.C.4.2.
III.C.4.2.
(p. 40)
(P. 47)
Page No. 8-25

-------
EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Multiple
Dilution Tube (MPN) Method
Page No. 8-26
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
5.	Processing discarded
cultures.
6.	Disinfection
5.	Return negative LLSTB cul-
tures to 35 C incubator.
6.	Place the separate rack of
newly inoculated EC Broth
tubes in water bath
incubator.
1.	Sterilize discarded LLSTB
tubes.
2.	Remove all labels from
culture tubes .
3.	Empty sterilized cultures
into sink.
4.	Wash and dry culture tubes,
fermentation vials, and
tube caps.
1. Disinfect laboratory bench
top; wipe dry.
5a. An additional 24 + 2 hours at 35 + 0.5°C.
5b. Rack should contaTn all LLSTB gas-negative tubes.
6a. 24+2 hours at 44.5 + 0.2°C.
6b. Must be put in incubator within 30 minutes after
transfers have been made.
la. Autoclave: 15 minutes at 121°C.
2a. Best done while still warm after autoclave.
4a. Meets original cleanliness requirements of
glassware.
4b. Tubes and caps ready for re-use.
la. Sponge and disinfectant; paper toweling.

D. 48-hour Procedures
1.	Equipment
Maintenance
2.	Disinfection
1.	Check, record, and adjust
incubator temperatures.
2.	Add water to pan in incu-
bator as necessary.
1. Disinfect lab bench top;
wipe dry.

-------
-------
EFFLUENT MONITORING PROCEDURE: Fecal Col 1form Test by the Multiple
Dilution Tube (MPN) Method
Page No. 8-28
DERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
i. Processing discarded
tubes of media.
5. Disinfection
5a. (alternate) If no cultures
remain to be returned to
incubator, proceed to
"Interpretation of Test
Results" and continue as
directed.
1.	Sterilize discarded media.
2.	Remove all labels from
culture tubes.
3.	Empty sterilized cultures
into sink.
4.	Wash and dry culture tubes,
fermentation vials, and
tube caps.
1. Disinfect laboratory bench
top; wipe dry.
E. 72-hour Procedures
1. Equipment
maintenance
2.	Disinfection
3.	Readfno and record-
ing i jresults.
1.	Check, record, and adjust
incubator temperatures.
2.	Add water to pan 1n Incu-
bator as necessary.
1. Disinfect lab bench top;
wipe dry.
1. Remove rack(s) of culture(s)
from water bath Incubator to
lab bench.
-------
FFFlDENT MONITORING PROCEDURE: Fe«l Coliform Test by the Multiple
—¦¦-¦"¦	Dilution Tube (MPN) Method
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
E. 72-hour Procedures



(Continued)




2. Shake culture rack(s)



gently.



3. Examine each tube for gas



production and record re-



sults on data sheet.



4. Place all tubes of EC



Broth 1n discard basket.


4. Processing discarded
1. Sterilize discarded tubes


tubes of media
of media.



2. Remove all labels from



tubes.



3. Empty sterilized tubes



into sink.


5. Disinfection
1. Disinfect lab bench top;



wipe dry.


F. Interpretation of
tsst results (Con-
tinued)
1.	Determine number of EC
Broth gas-positive tubes
for each group of five
tubes of equal sample
volumes.
2.	Write the numbers in the
data sheet.
la. Assume, for instructional purposes,
5 positive 1st row
5 positive 2nd row
2 positive 3rd row
0 positive 4th row
II-F.I
(P. 37)
II.F.2
(P- 37)
Page No. 8-29
-------
EFFLUENTJ10NIT0KIN6 PROCEDURE: Fecal Coliform Test by the Multiple
Dilution Tube (MPN) Method
Page No. 8-30
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
F. Interpretation of
test results (Con-
tinued)
-------
EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Multiple
Dilution Tube (MPN) Method
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
F. Interpretation of
test results (Con-
tinued)
3. Select the 3-digit code
which applies to the
number of gas-positive
tubes of EC Broth.
4. Look up and record on the
data sheet the WN Index.
3a. In a test involving 4 sample volumes this will
be based on rows 1, 2, 3, or on rows 2, 3, 4;
and
3b. If all tubes are positive in rows 1 and 2, then
the 3-digit code is based on rows 2, 3, 4.
3c. In all other cases the 3-digit code is based on
rows 1,2,3.
4a. For the given example the location of the MPN
index is shown by the arrow based on the 5-2-0
code.
II.F.3
(p. 37)
(P. 38)
No. of Tubes Giving Positive
MPN
Reaction out of

Index



per



100 ml
5 of 10
5 of 1
5 of 0.1

ml Each
ml Each
ml Each

5
1
0
33
5
1
1
46
5
1
2
63
-*•5
2
0
49
5
2
1
70
A
A

A
II.F.4
(P. 32)
II.F.5
(P- 38)
Page No. 8-3.1
-------
EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Multiple
¦——1———1— Dilution Tube (MPN) Method
Page No. 8-32
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
F. Interpretation of
test results (Con-
tinued)
5.	Divide the MPN Index by
the number of ml of sample
represented by the middle
diqit of the MPN Code.
The number obtained is the
MPN (Most Probable Number)
per 100 ml of original
sample.
6.	Record the calculated
Total Conforms per 100 ml
on the laboratory data
sheet.

RESULTS:
Fecal Coliform MPN
4900

II.F.6
(p. 38)
G. Reporting of results
1. Report results as pre-
scribed under NPDES or
other regulatory require-
ments.
la. Report Geometric Mean
lb. See procedure for calculating Geometric Mean
described elsewhere in these instructions (EMP
units).





-------
Effluent Monitoring Procedure: Fecal Coliform Test by the Multiple
Dilution Tube (MPN) Method
TRAINING GUIDE
SECTION	TOPIC
I*	Introduction
II*	Educational Concepts - Mathematics
III*	Educational Concepts - Science
IV	Educational Concepts - Communications
V*	Field & Laboratory Equipment
VI*	Field & Laboratory Reagents
VII*	Field & Laboratory Analyses
VIII	Safety
IX	Records and Reports
~Training guide materials are presented here under the headings marked *.
These standardized headings are used through this series of procedures.
Page No. 8-33
-------
EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Multiple
Dilution Tube (MPN) Method
INTRODUCTION	Section I

TRAINING GUIDE NOTE
REFERENCES/RESOURCES
B.6.1.1
These MPN methods for determining bacterial numbers
are based on the assumption that the bacteria can
be separated from one another (by shaking or other
means) resulting in a suspension of individual
bacterial cells, uniformly distributed through the
original sample when the primary inoculation is
made.
Test procedures are based on certain fundamental
assumptions:
a.	First, even if only one living cell of the
test organisms is present in the sample, it
will be able to grow when introduced into the
primary inoculation medium;
b.	Second, growth of the test organism in the
culture medium will produce a result which
indicates presence of the test organism; and;
c.	Third, unwanted organisms will not grow, or
if they do grow, they will not limit growth
of the test organism; nor will they produce
growth effects that will be confused with those
of the bacterial group for which the test is
designed.

Page No. 8-34
-------
EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Multiple
Dilution Tube (MPN) Method
EDUCATIONAL
CONCEPTS
- MATHEMATICS

SECTION II

TRAINING
GUIDE NOTE

REFERENCES/RESOURCES

Table of Most Probable Numbers (MPN)


No of Tubes Giving Positive
MPN


Reaction out of

Index





per





100 ml


5 of 10
5 of 1
5 of 0.1



ml Each
ml Each
ml Each



0
0
0
<2


0
0
1
2


0
1
0
2


0
2
0
4


1
0
0
2


1
0
1
4


1
1
0
4


1
1
1
6


1
2
0
6


2
0
0
5


2
0
1
7


2
1
0
7


2
1
1
9


2
2
0
9


2
3
0
12


3
0
0
8


3
0
1
11


3
1
0
11


3
1
1
14


3
2
0
14


3
2
1
17



3
0
17


4
0
0
13


4
0
1
17


4
1
0
17


4
1
1
21


4
1
2
26


4
2
0
22


4
2
1
26


4
3
0
27


4
3
1
33


4
4
0
34


5
0
0
23


5
0
1
31


5
0
2
43

Page No. 8-35
-------
EFFLUENT MONITORING PROCEDURE:
Fecal Coliform Test by the Multiple
Dilution Tube (MPN) Method
EDUCATIONAL CONCEPTS - MATHEMATICS
SECTION II
TRAINING GUIDE NOTE
REFERENCES/RESOlir.ClS
Table of Most Probable Numbers (MPN)
No. of Tubes Giving Positive
Reaction out of
5 of 10
ml Each
5
5
5
5
5
5
5
5
5
5
5
5
5
5
5
5
5
5
5
5
5
5 of 1
ml Each
2
2
2
3
3
3
3
4
4
4
4
4
5
5
5
5
5
5
5 of 0.1
ml Each
0
1
2
0
1
2
0
1
2
3
0
1
2
3
4
0
1
2
3
4
5
MPN
Index
per
100 ml
33
46
63
49
70
94
79
110
140
180
130
170
220
280
350
240
350
540
920
1600
!2400
Page No. 8-36
-------
EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Multiple
Dilution Tube (MPN) Method
EDUCATIONAL CONCEPTS - MATHEMATICS
Section II
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
F.l
F.2
F.3
For purely qualitative aspects of testing for
indicator organisms, it is convenient to consider
the tests applied to one sample portion, inoculated
into a tube of culture medium, and the follow-up
examinations and tests on results of the original
inoculation. Results of testing procedures are
definite: positive (presence of the organism-group
is demonstrated or negative (presence of the or-
ganism-group is not demonstrated).
The combination of positive and negative results
is used in an application of probability mathe-
matics to secure a single MPN value for the sample.
To obtain MPN values, the following conditions
must be met:
a.	The testing procedure must result in one or
more tubes in which the test organism Is demon-
strated to be present; and
b.	The testing procedure must result in one or
more tubes in which the test organism is not
demonstrated to be present.
The MPN value for a given sample is obtained
through the use of MPN Tables. It is emphasized
that the precision of an individual MPN value is
not great when compared with most physical or
chemical determinations.
Standard practice 1n water tests made by this or-
ganization Is to plant five tubes in each of a
series of sample increments, in sample volumes
decreasing at decimal intervals.
As an example, assume that all tubes were positive
for a sample portion of 1.0 ml, all five tubes
were positive on the portions of 0.1 ml, two of
the five 0.01 ml portions were positive, and none
of the five 0.001 ml portions were positive.
1. The numbers, on the above example, would be
5-5-2-0.
1. Pursuing the above example, the code would be
5-2-0.
Page No. 8-37
-------
EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Multiple
Dilution Tube (MPN) Method
EDUCATIONAL CONCEPTS - MATHEMATICS
Section II
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
F.3 (Continued
F. 4
F. 5
F. 6
2. Selection of codes is sometimes complicated.
For further information study training guide
notes and cited references.
1.	Appears on MPN Table (attached to this Section)
2.	Pursuing the above example, the MPN Index for
MPN Code 5-2-0 would be 49.
1. As indicated above, the middle digit is 2; and
it represents a sample portion of 0.01 ml. An
MPN Index of 49 divided by 0.01 is 4900.
The Fecal Coliforms per 100 ml would be recorded
as 4900.
Std. Meth. 14-923 ff
Page No. 8-
38
-------
EFFLUENT MONITORING PROCEDURE: Fecal Coll form Test by the Multiple
Dilution Tube (MPN) Method
EDUCATIONAL CONCEPTS - SCIENCE
Section ni
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
C.3.3
Interpretation of results on LLSTB:
Development of gas in this medium indicates that
the lactose has been fermented. Fermentation of
lactose with gas production is a basic characteris-
tic of coliform bacteria. To meet the defination
of coliforms, gas must be produced from lactose
within 48 hours after being placed in the incubator.
If a culture develops gas only after more than 48
hours incubation, then, by definition, it is not a
coliform.
Meeting previously discussed assumptions (See
B.6.1.1) usually makes it necessary to conduct the
tests in a series of stages.
Features of a full, multi-stage test:
a. First stage: The culture medium usually serves
primarily as an enrichment medium for the group
tested. A good first-stage growth medium should
support growth of alj_ the living cells of the
group tested, and it should include provision
for indicating the presence of the test organism
being studied. A first-stage medium may include
some component which inhibits growth of ex-
traneous bacteria, but this feature never should
be included if it also inhibits growth of any
cells of the group for which the test is de-
signed. The Presumptive Test for the coliform
group is a good example. The medium supports
growth, presumably, of all living cells of the
coliform group; the culture container has a
fermentation vial for demonstration of gas pro-
duction resulting from lactose fermentation by
coliform bacteria, if present; and sodium lauryl
sulfate may be included in one of the approved
media for suppression of growth of certain non
coliform bacteria. This additive apparently has
no adverse effect on growth of members of the
coliform group in the concentrations used. If
the result of the first-stage test is negative,
the study of the culture is terminated, and the
result is recorded as a negative test. No fur-
ther study is made of negative tests. If the
result of the first-stage test is positive, the
culture may be subjected to further stuc(y to
verify the findings of the first stage.
Page No. 8-39
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EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Multiple
Dilution Tube (MPN) Method
EDUCATIONAL CONCEPTS - SCIENCE	Section in

TRAINING GUIDE NOTE
REFERENCES/RESOURCES
C.4.2
Transfer of gas-positive LLSTB Tubes to EC Broth:
This is done in order to find out if the organisms
which produced gas from the lactose in LLSTB also
can produce gas from a slightly different culture
medium (it also contains lactose), and can do so
at an elevated temperature (44.5 + 0.2 C.) in a
water bath. Practically all coliforms which came
from intestinal wastes are able to produce gas from
lactose at the elevated temperature of the second
medium; and practically all bacteria which produce
gas from lactose, but which do not come directly
from intestinal wastes, are unable to perform at
elevated temperature.

Page No. 8-40
-------
EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Multiple
Dilution Tube (MPN) Method
FIELD AND LABORATORY EQUIPMENT AND SUPPLIES	Section V

TRAINING GUIDE NOTE
REFERENCES/RESOURCES
A.1.1
Incubator should be kept out of drafts or direct
sunlight in order to prevent temperature inside
the incubator from changing outside the tempera-
turn range specified (35 +0.5 ).
Power supply should be selected so that there won't
be too many pieces of-equipment on the same cir-
cuit. Otherwise, circuits will be blown repeatedly.
Standard Methods for the
Examination of Water and
Wastewater, Nth ed. f 1975)
APHA, WPCF, AWWA, p. 880
(Hereafter referred to as:
Std. Meth. 14: (page no.)
A.1.2
Mercury bulb thermometer usually used in most incu-
bators. Recording thermometer is acceptable, but,
it should be calibrated against a mercury bulb
thermometer which has been certified by National
Bureau of Standards. The NBS certified thermo-
meter always should be used with its certificate
and correction chart.

A.1.3
Saturated relative humidity is required in order
to make the incubation more efficient (heat is
transferred to cultures faster than in a dry incu-
bator). Furthermore, culture medium may evaporate
too fast in a dry incubator.

A.1.5
Allow enough time after each readjustment to per-
mit the incubator to stabilize before making a
new adjustment. At least one hour is suggested.

A.1.6
Incubator temperature can be held to much closer
adjustment if operated continuously. Temperature
records should be kept in some form of permanent
record. A temperature record book is suggested.
If a recording thermometer is used, the charts may
be kept as permanent record; if so, be sure that
the charts are properly labeled to identify the
Incubator and the period covered.

A.3.1-5
Since electric sterilizer will be operated inter-
mittently, care should be taken that it is on a
circuit which will not be overloaded when it is
turned on.
Std. Meth. 14:881
A.4.1
Autoclaves differ greatly 1n design and in method
of operation. Some are almost like home-style
pressure cookers; others are almost fully auto-
matic. This 1s a subject which requires separate
Instruction; and should be related to the exact
make and model of equipment you will use in your
own laboratory.
Std. Meth. 14:881
Page No. 8-41
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EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Multiple
Dilution Tube (MPN) Method
Section

TRAINING GUIDE NOTE
REFERENCES/RESOURCES
A.5.1-2
Distilled water in a bacteriological laboratory
must not contain substances which will prevent
any bacteria from growing in culture medium in
which the distilled water is used or will be highly
nutritive. There are procedures for testing qual-
ity of distilled water; but these should be under-
taken only by professional bacteriologists or in
laboratories where this is done regularly. Use
only glass stills or block tin lined stills.
Std. Meth. 14:645-49
14:888-891
Training Manual (EPA)
Current Practices in Water
Microbiology
A.6.1
pH Meter: see cited reference
Std. Meth. 14:882
A.7.1-4a
Glassware: See cited reference on pi pets and
graduated cylinders, media utensils, bottles.
Std. Meth. 14:882-885
A. 7.1-4b
Glassware can be checked for bacteriostatic or in-
hibitory residues by a bacteriological test pro-
cedure which, like the distilled water suitability
test, should be undertaken only by professional
bacteriologists or in laboratories where this test
is done on a regular basis.

A.10.1-6
Sample bottles:
Wide-mouthed glass-stoppered bottles suggested,
but other styles acceptable.
If glass-stoppered bottles are used, a strip of
paper should be placed in the neck of the bottle
before placing the stopper in place in preparation
for sterilization. This prevents the glass stopper
from "freezing" in place during sterilization. The
paper strip is discarded at the time of sample
collection.
Std. Meth. 14:884
14:904
A.11.1-6
Pi pets:
This procedure is described in terms of reusable
glass pipets. However, single-service prepackaged
glass or plastic pipets may be purchased and used,
if preferred. In case of use of single-service
pipets, they will be sterile when purchased, are
used one time, and discarded immediately after use.
Accordingly, in the step-by-step procedures dis-
regard any instructions about preparation of pipets
for reuse in case of using single-service pipets.
Std. Meth. 14:882-883
Page No. 8-42
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AFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Multiple
Dilution Tube (MPN) Method
FIELD AND LABORATORY EQUIPMENT AND SUPPLIES

TRAINING GUIDE NOTE
REFERENCES/RESOURCES
A.11.7
Passing the opened can of pi pets through a flame
burns off excess cotton wisps sticking out of the
mouthpiece of the pipet. If this is not done, it
is almost impossible to control sample measurement
accurately.

A. 12.Id.
See cited reference. In time, this solution will
become mold-infested. At this time it should be
discarded and a new stock solution prepared.
Std. Meth. 14:892
A.12.3
Dilution water preparation:
Measurement of dilution water into bottle with a
100 ml graduated cylinder is time-consuming, but
effective. An automatic pipetting machine can be
considered a luxury, but is a real time-saver.

A.12.4
If caps are not placed on bottles of dilution water
loosely, they may crack in autoclave; furthermore,
steam will not be able to get in contact with the
material being sterilized. After sterilization,
tightening caps on bottles of distilled water will
permit them to be kept for long periods.

A.12.5
Always pack material loosely and away from walls
in autoclave when preparing to sterilize. Steam
must flow freely around materials being sterilized.

A.12.7
If water should evaporate noticeably or become
contaminated by microbial growth, the bottle of
distilled water should be discarded.

A.13.3b
Funnel \
Nw
L£
Funnel, Hose, and
—y Pinchcock Assembly
Pi nchcock
v-— Hose
¦»_ Glass Tube
NOTE: Unit need not be
sterile for medium
delivery only

Page No. 8-43
-------
EFFLUENT MONITORING PROCEDURE: Fecal Col 1 form Test by the Multiple
Dilution Tube (MPN) Method
FIELD AND LABORATORY ANALYSES
SECTION VTT
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
There is no such thing as a "standard" data sheet
for bacteriological tests. A simplified data sheet
is shown below:
In this procedure, it is recommended that the worker
learn to select a series of 4 sample volumes in de-
creasing amounts as indicated.
It is possible to use as few as three sample volumes,
but often the worker will fail to get a measurable
result. On the other hand, one could have 5, 6, or
even more sample volumes in decreasing amounts.
Fecal Coliform Test
Multiple Dilution Tube (MPN) Method
Sample Type ____	Lab. No.
Station	Description	
Collection Date	Time	APM. Temp. 	
„ . AM. AM
Received 	 PM. Examined 	 PM.
pH 	Observations
Amount
Sample
Presumtlve
LST
Fecal
EC
No.
Pos
EC




m

24 hr
48 hr
24 hr

































































'















































































































Page No. 8-44
Results:
Fecal
coliform MPN
\
-------
EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Multiple
Dilution Tube (MPN) Method
FIELD AND LABORATORY ANALYSES
SECTION VII
TRAINING GUIDE MOTE
REFERENCES/RESOURCES
Suggested labeling code for tubes:
1.	Every tube shows the "laboratory bench number
(323 in example shown on sample data sheet).
2.	Below the laboratory bench number on each tube
will be found a coded symbol which represents the
sample volume and the tube of each series of five,
Thus:
Sample volume, ml Tubes are labeled
1.0
O.T
0.01
0.001
0.0001
0.00001
0.000001
0.0000001
a, b, d, d,	e
a, b, c, d_,	e,
Ta,~lb, lc,	Td,	le
2a, 2b, 2c,	2d,	2e
3a, 3b, 3c,	3d,	3e
4a, 4b, 4c,	4d,	4e
5a, 5b, 5c,	5d,	5e
6a, 6b, 6c,	6d,	6e
etc., etc,
3.
For example, a tube might look something like
this, to represent sample No. 323, with the •
middle tube of a series of five, representing
0.1 ml:
[3231
C
Paae No. 8-45
-------
EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Multiple
Dilution Tube (MPN) Method
FIELD AND LABORATORY ANALYSES
Section VII
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
B.6.2-3
Multiple dilution tube tests for quantitative de-
terminations apply a Most Probable Number (MPN)
technique. In this procedure one or more measured
portions of each of a series of decreasing sample
volumes is inoculated into the first-stage culture
medium. Through decreasing the sample increments,
eventually a volume is reached where only one cell
is introduced into some tubes. Each of the several
tubes of sample-inoculated first-stage medium is
tested independently, according to the principles
described.
Sample dilutions and inoculations: See Figure 2 as
another way to represent sample dilution and inocu-
lation. Note that sample dilutions are made as
needed durinq the inoculation procedure; they are
not made up before starting to inoculate tubes of
culture medium. Bacteria shall not be suspended
in any dilution water for more than 30 minutes at
room temperature.
Dilution Ratio*:
Figure 2. PREPARATION OF DILUTIONS
1:100
1:10000
99 ml
blank
1 ml
1 ml
Delivery volume 1 ml
0/1 ml
1 ml
0.1 ml
Water
Sample
Delivery volume 1 ml

1 ml
0.1 ml
Tubes
Actual volume
of sample In tube
1 ml
Petri Dishes or Culture Tubes
0.1 ml 0.01 ml 0.001 ml
0.0001 ml 0.00001 ml
Page No. 8-46
-------
EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Multiple
Dilution Tube (MPN) Method
FIELD & LABORATORY ANALYSES
Section VII
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
C.4.2
Transfers of LLSTB
Transfers can be made, as indicated, with a wire
loop having a diameter of at least 3 mm. An
alternate method of transfer authorizes the use of
an "applicator stick" which is a single service
hardwood transfer device. Its dimensions are 0.2
to 0.3 cm in diameter and 2.5 cm longer than the
test tube used in the analysis. The term single
service denotes that the stick is pre-sterilized
and used for a single transfer (LLSTB to EC) and
then discarded in the pan containing disinfectant
and a new sterile stick used for the next tube to
be transferred. Use of this stick technique makes
the gas burner unnecessary for the transfer process,
Std. Meth. 14:922
Page No. 8-47
-------
A PROTOTYPE FOR DEVELOPMENT OF
ROUTINE OPERATIONAL PROCEDURES
for the
FECAL COLIFORM TEST
by the
MEMBRANE FILTER METHOD
as applied in
WASTEWATER TREATMENT FACILITIES
and in the
MONITORING OF EFFLUENT WASTEWATERS
Developed by the
National Training and Operational Technology Center
Municipal Operations and Training Division
Office of Water Program Operations
U.S. Environmental Protection Agency
W.BA.EMP.3a.6.77
Page No. 9-1
-------
EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Membrane
Filter Method
This Procedure was developed by:
NAME	Rocco Russomanno
ADDRESS EPA, OWPO, NTOTC, Cincinnati, Ohio 45268
POSITION Microbiologist-Instructor
EDUCATION AND TECHNICAL BACKGROUND
B.S. - University of Massachusetts
5	years Envorcement Action, Raritan Bay Project, PHS & DI
10 years Instructor,
NAME	Harold L. Jeter
ADDRESS EPA, OWPO, NTOTC, Cincinnati, Ohio 45268
POSITION Chief, Program Support Training Branch
EDUCATION AND TECHNICAL BACKGROUND
A.B. - James Millikin University
M.A. - Ohio University
4 years - US Army; Clinical Laboratories; specializing
bacteriology
3 years College Instructor - Bacteriology
6	years Research in Sanitary Microbiology
22 years Training of Federal, State and Local personnel 1n
principles and practice of sanitary bacteriology of water
Page No,
-------
Page No. 9-4
Fecal Coliform Test
Membrane Filter Method
Flow Sheet

M-FC Medium
Sam
(3 sample volumes filtered:
20 ml, 10ml, and 5 ml)
hi
1
[20
(Similar to rest
of flow sheet)
[10 ml]
Incubate fori 24 hours at
44.5°C + 0.2°C in a constant
temperature water bath
[ 5 ml]
(Similar to rest
of flow sheet)
Colonies counted microscopically
(10 to 15 magnifications)
Fecal Coil forms
(Colonies with blue or
blue-tinted colorations)
Obtain number of colonies
Calculate fecal coliforms/100 ml
I	
Record
Non-Fecal Coliforms
(Colonies lacking blue
or blue-tinted colorations)
Fecal coliforms
Not present
Results as fecal coliforms/100 ml
Report results as prescribed under NPDES
or other regulatory requirements
-------
EFFLUENT MONITORING PROCEDURE: Membrane Filter Test Method
1.	In wastewater effluent quality control, the application of this methodology
can be for one or both of the following;
a.	To determine whether the bacteriological quality of the effluent meets
quality requirements set by law or by regulatory authority; and,
b.	To determine the bacteriological effects of effluent water on the
bacteriological quality of the receiving water.
2.	Brief description of analysis:
A series of measured sample portions is filtered through membrane filters
placed individually within a filtering apparatus. Bacteria in the samples
are held on the upper surfaces of the filters, while the water passes
through and is discarded.
The membrane filters are placed on a special culture medium, called M-FC
Broth, in plastic petri dishes. The inverted petri dishes are placed in
a leakproof plastic bag, and incubated totally immersed in a water bath
at 44.5 + 0.2 C for 24 hours + 2 hours. On M-FC Broth, fecal coliform
will grow and develop blue or blue-tinted colonies. Colonies lacking this
color characteristic are not considered as fecal coliforms. The blue color
may appear only in the centers of the colonies, or entire colonies may be
colored. Very few other colonies will develop on the medium at the stated
incubation temperature.
One or two membranes are selected for colony counting on the basis of
suitable colony density, and colonies are counted with the aid of a
binocular dissecting microscope at a magnification of 10X or 15X. After
colonies are counted, a calculation is made in order to report fecal
coliforms per 100 ml.
3.	ApplicaDi1ity of this Procedure:
a.	Range of Concentration:
This procedure, as outlined, will detect fecal coliforms within the range
of 100 to 1200.
b.	Pretreatment of Samples:
In accordance with Standard Methods, 14th ed. (p. 904) and as outlined in
EMP, "Collection and Handling of Bacteriological Samples."
Analytical Method: Standard Methods for the Examination of Water and Wastewater.
14th ed., 1975, pg. 937 ff.
Page No. 9-5
-------
EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Membrane
FiIter Method
General description of equipment and supplies used in the test analysis
A.	Capital Equipment
Autoclave, steam - providing unifrom temperatures up to and including 121°C
and equipped with an accurate thermometer, pressure gauges, saturated
steam power lines and capable of reaching required temperatures within
30 minutes.
Balance - Sensitivity of 0.1 gram at a load of 150 grams, with appropriate
weights.
Incubator, waterbath - having forced circulation and provided with aQcover.
Must be capable of providing an incubation temperature of 44.5 + 0.2 C.
Oven, hot-air- providing unifrom temperatures within the range of 160 - 180°C.
Meter, pH - accurate to within 0.1 pH unit, with suitable standard pH re-
ference solution (s).
Apparatus, water distillation - suitable for bacteriological culture media
(alternately, a suitable source is permissible).
Microscope, sterioscopic - 10X - 15X magnification with fluorescent lighting
preferred. Alternately, a small fluorescent lamp with magnifier is
acceptable.
Refrigerator - Set for less than 10°C but above the freezing temperature. If
sample cannot be run within 1 hour refrigeration will be necessary.
Vacuum Source - preferably a pump assembly with suitable hoses and shut-off
clamp or valve provided. As an alternate method an aspirator or hand pump
with the same provisions are acceptable.
Filtration Unit, MF - a seamless funnel attached to a receptacle bearing a
porous plate (screen, porous disc, etc.), stainless steel, glass, porcelain
or other suitable material.
B.	Reusable Supplies
Apron - suitable for laboratory operations.
Bottle, sample - 250 ml, wide-mouth, glass stopper, with tag (used for
sampling operations).
Bottle, squeeze type - containing disinfecting solution.
Burner, gas - suitable for laboratory operations
Can, pipet - non-toxic and sterilizable material (if pre-sterilized dis-
posable-type pi pets are used, this item is unnecessary).
Pan, discard - receives contaminated pipets.
Graduate cylinder 100 ml; 500 ml
Paqe No. 9-6
-------
Pi pets, microbiological - 1.0 ml, with 0.1 ml graduations, sterile cotton
plugged, glass or disposable types (the disposable types are for one time
use and may be glass or plastic).
Pi pets, microbiological - 10 ml, with 1 ml graduations, sterile, cotton-
plugged, glass or disposable types (the disposable types are for one time
use and may be glass or plastic).
Thermometer (water batb) - must indicate within the 40° - 50°C range and
have increments of 0.1 C, NBS (National Bureau of Standards) or calibrated
against NBS thermometer. Full immersion type preferred.
Thermometer (oven) - must indicate within the 160 - 180°C range and have
increments at least 1.0 C.
Glassware, borosilicate
beaker, 50 ml (for measuring pH, rosolic acid
Flask, volumetric, 1 liter capacity (for stock solution of phosphate buffer)
Flask, Erlenmeyer, 500 ml capacity (for holding buffered distilled rinse water)
Flask, sidearm 1 liter size (for reservoir of MF apparatus. Proper sized and
bored rubber stopper is needed to connect MF filtration flask to unit).
Flask, Erlenmeyer, 250 ml (for preparing MFC medium)
Forceps, curved end, round tip
Bottle, small, Methanol or Ethanol volume to cover ends of forceps
Sponge, small, to spread and wipe germicide
Desiccator, media storage. Ideally opaque or darkened and containing
desiccating agent to remove moisture.
C. Consumable Supplies:
Dish, petri, disposable, tight fitting plastic, 50 x 12 mm, sterile.
M-FC Broth medium dehydrated, fecal coliform. Distributors Difco, BBL or
other equivalent preparation.
Rosolic Acid reagent, 50 gram bottle, Allied Chemical, Olin Matheson
Difco or equivalent preparation.
Filter, membrane, 47 mm, 0.45 pm pore size, white, grid marked, sterile.
Pad, absorbent 48 mm, sterile (usually included with membrane packet)
Bag, plastic, water-proof, closure provided or method of sealing bag
necessary for water immersion.
Page No. 9-7
-------
Disinfectant, dilute iodine aqueous (water) solution. Commercial preparation
or 1 gram iodine crystals and 2 grams potassium iodide to a liter of distilled
water.
Methanol or Ethanol, absolute, (for forceps disinfection)
Water, distilled, buffered, sterile (for MF funnel rinsing)
Stock solution, buffer, potassium dihydrogen phosphate
Water, distilled, suitable for bacteriological operations
Potassium Dihydrogen Phosphate (KH^PO^) reagent, 1 lb. unit
Data sheet suitable for fecal coliform procedures (has pertinent field
information [location, time, sampler, etc.]; lab information [sample,
mis filtered, colony counts, etc.]; and effluent monitoring required
data [fecal coliforms/100 ml]).
D. Expendable Laboratory Supplies:
Marker, glass or plastic
Glass Wool
Non-absorbent Cotton
Paper, kraft wrapping
Tape, autoclave pressure-resistant
Foil, aluminum, heavy duty
Matches or striker
Toweling, paper
Item needs in quantities or required size or space allowances cannot be specified,
as they vary according to the daily analysis schedule. As a rule-of-thumb,
space/size or quantity requirements should be at least 3 times the normal daily
requirements. For further information on specifications for equipment and supplies,
see the Microbiology Section of the current edition of "Standard Methods for the
Examination of Water and Wastewater."
Page No. 9-8
-------
EFFLUENT MONITORING PROCEDURE: Feca1 Coliform Test by the Membrane Filter Method
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
A. Pretest Procedures
1. Water bath incuba-
tor setup,
adjustment (44.5 C
+ 0.2 C.)
1. Place water bath incubator
in permanent location.
2. Put water in water bath.
3. Install thermometer.
4.	Connect water bath incu-
bator to electric power
source and turn on.
5.	Adjust temperature until
stabilized at required
temperature.
6. Operate water bath incu-
bator continuously.
Aa. All pretest procedures completed before starting
other first-day procedures
la. On bench or table surface.
lb. Out of drafts or place in which it will be in
direct sunlight part of day.
lc. Location convenient to laboratory bench.
Id. Convenient source of electric power, separate
circuit is possible.
2a. Distilled or deionized water preferred, tap water
accepted.
2b. Should be deep enough to permit total immersion
of the plastic bags containing petri dishes.
Usually this is about 2% -3 inches above the
platform in the waterbath.
3a. Functions at least in 40° - 50°C range. Meets
NBS standards. Have at least 0.1 C increment
markings.
3b. Most water baths provide for corner location for
thermometer (for protection from breakage).
4a. Pilot light should come on.
5a. Manufacturer's instructions for location and
method of temperature adjustment.
5b. Allow about 1 hour between adjustments.
5c. Operation must be at 44.5 +0.2 C.
6a. Requires daily check with written temperature
record, with adjustment as necessary.
6b. Requires daily check of water level and addition
of more as needed.
6c. With tap water in water bath, may require
periodic scum removal from inner walls.
V.A.1.1
V.A.1.2
V.A.1.3
Page No. 9-9
-------
EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Membrane Filter Method
Paqe No. 9-10
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
2. Oven sterilizer
setup
1.
Place oven sterilizer in
permanent position.
la. A convenient source of electric power.


2.
Install thermometer
2a. Should read in the 160-180°C range, be accurate
within this interval, and be marked in 1.0 degree
intervals.


3.
Connect sterilizer to
power source and turn ON.
3a. Pilot light or element heating effect indicates
power ON.


4.
Adjust oven temperature to
stabilize at required
sterilizing temperature.
4a. 170°C is required temperature.


5.
Operate when sterilizing
is required.
5a. Turned ON in advance of use and checked for
temperature stabilization.
5b. Used for dry glassware and metal objects which
can be covered by a paper or metallic foil
covering.
5c. Not used for culture media, liquids, plastics,
and rubber objects or anything containing or
including these.

3. Autoclave setup
1.
Install and operate auto-
clave according to manufac
turer's instructions.
la. Variable in design and operation, and unless
properly operated can be dangerous,
lb. Used to sterilize objects made of or including
liquids, rubber, and some plastics and for
glassware, if desired,
lc. Operated for general sterilization at 121 C
(250 F) for a period of 15 minutes after this
temperature has been attained.
Id. Sterilized media and liquids must be removed as
soon as possible upon sterilization.
V.A.3
-------
EFFLUENT MONITORING PROCEDURE: Feca1 Co1lform Test by the
OPERATING PROCEDURES
STEP SEQUENCE
4. Water distillation
equipment
5. pH meter
6. Glassware
1.	Install and operate in
accordance with manu-
facturer's instructions.
2.	Operate as required to
maintain adequate supplies
of distilled water.
1. Setup and operate in
accordance with manufact-
urer's recommendations.
1.	Cleaned and rinsed using a
suitable detergent and
hot water.
2.	Use a final rinse of
deionized or distilled
water.
Membrane Filter Method
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
la. Must produce water meeting quality requirements
for bacteriological tests.
V.A.4
la. Meter must be accurate to at least 0.1 pH unit.
la. Non-toxic detergent must be completely removed
from glassware.
2a. 6 to 12 successive rinsings may be required.
2b. Must produce a dry glassware which meets
bacteriological requirements for suitability.
~IMPORTANT*
The following Special conditions may apply to the
sample to be analyzed:
*If sample is chlorinated effluent which contains
copper, zinc, or heavy metals do operating
procedures A.7, A.8, and A.9 completely.
*	If sample is unchlorinated efflu&nt which contains
copper, zinc, or heavy metals, eliminate steps:
A.7 and A.9.1
*If the sample is chlorinated effluent which does
not contain copper, zinc, or heavy metals, eliminate
steps: A.8 and A.9.2
*	If the sample is unchlorinated and contains no
aTa.-	e,,""nate ste»s
V.A.6.2
Page No. 9-11
-------
EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Membrane Filter Method
Page No. 9-12
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
7. Sodium thiosulfate
solution
Ethylene-
d i ni tri1otetra-acetic
acid (EDTA) solution
9. Sample bottle pre-
paration
1.	Weigh 10.0 grams of sodium
thiosulfate.
2.	Dissolve in 50-60 ml dis-
tilled water.
3.	Add distilled water to
bring final volume to
100 ml.
4.	Transfer to labeled bottle
1. Weigh 15.0 grams of EDTA.
2.	Dissolve in 50-60 ml of
distilled water.
3.	Add distilled water to
bring final volume to
100 ml.
4.	Transfer to clean labeled
bottle.
1.	Deliver 0.1 ml or 0.2 ml
of 102 sodium thiosulfate
solution to each sample
bottle. (.1ml to 4 oz or
120 ml size and .2 ml to
6-8 oz or 250 ml size)
2.	Deliver .3 ml or .6 ml of
15% EDTA solution to each
sample bottle (.3 ml to
4 oz or 120 ml size and
.6 ml to 6-8 oz 250 ml size
la. Used for dechlorination of smaples
lb. Use of trip balance accepted.
2a. 100 ml graduated cylinder satisfactory.
4a. Should be labeled as 10% sodium thiosulfate and
stored in refrigerator.
la. Used for water samples high in copper or zinc or
wastewater high in heavy metals,
lb. Use is trip balance accepted.
2a. 100 ml graduated cylinder satisfactory.
4a. Labeled as 15% Ethylene-dinitrilotetra-acetic
acid (EDTA) and stroed in refrigerator.
la. Use 1 ml pi pet
lb. Provides adequate sodium thiosulfate for
neutralizing chlorine in sample.
2a. Use 1 ml pi pet.
2b. Provides adequate EDTA chelating agent for metals
in sample.
2c. Return stock solution of EDTA to refrigerator.
V.A.9.1-6
-------
EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Membrane Filter Method
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
9. Sample bottle pre-
paration (continued)
3.
Place cover on sample
bottle.



4.
Place paper or metal foil
cover over bottle cap or
stopper.
4a. Protects opening of sample bottle from
accidental contamination.


5.
Sterilize sample bottles
in steriiizing oven.
5a. 1 hour at 170°C.


6.
Store sample bottles in
clean, dry place until
used.


10. Pi pets
1.
Insert a plug of non-
absorbent cotton into
mouthpiece of clean, dry
pi pet.
la. Pipets which have chipped or broken tips or tops
should be discarded,
lb. Cleanliness of pipet must be equivalent to glass-
ware.
lc. Non-absorbent cotton plug must be tight enough to
prevent easy removal, either by the pipeting
action or by handling, and yet be loose enough
to permit easy air movement through the plug.
Id. Plug protects user from ingesting sample into
his mouth.
V.A.10.1-5

2.
Pass plugged end of pi pet
quickly through burner.
2a. Removes wisps of cotton which prevent fingertip
control of pipeting action.


3.
Insert a layer of glass
wool or multi-layer of
paper padding in bottom of
pipet can.
3a. This protects tips from breakage.


4.
Place pipet in pipet can
with delivery tip down-
ward.
4a. Cotton-plugged end is pipeting end and opposite
end is delivery tip.

Page No. 9-T3
-------
EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Membrane Filter Method
Paqe No. 9-14
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
10.	Pi pets (continued)
11.	Blanks, dilution
water
5.	Sterilize pi pets in oven
or autoclave.
6.	Store cans in clean dry
place until needed.
1.	Prepare stock solution of
potassium dihydrogen phos-
phate (KH^PO^) by dissolv-
c$?0 grams of this
chemical in 500 ml of
distilled water and adjust
ing its pH to 7.2 with
IN NaOH. Dilute to 1
liter in a volumetric
flask.
2.	Prepare stock solution of
magnesium sulfate
(MgSO^HgO) by dissolving
50 grams of this chemical
in 500-600 mis of dis-
tilled water and, after
complete dissolving, bring
the final volume to 1 litei
| in a volumetric flask.
4b. Approximately 20 1 ml pipets or 12 ml pipets
will normally be accommodated in these cans.
4c. Can must be able to withstand steam pressure and
dry heat. Toxic materials, such as copper, are
not to be used. Aluminum is acceptable.
5a. At least 1 hour in oven at 170°C, or
in autoclave for 15 minutes at 121 C (autoclave
set for quick venting of steam).
5b. Cans removed quickly from autoclave with the aid
of asbestos gloves.
5c. Cans opened slightly to allow residual steam to
escape for a few seconds and then close can.
la. Distilled water may be measured in 500 ml gradu-
ated cylinder.
lb. Label to show contents, identity of preparer,
and date of preparation.
lc. Stored in refrigerator.
Id. Discarded if mold or turbidity appears.

-------
EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Membrane Filter Method
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES

3.	Prepare workinq solution
of dilution water by
adding 1.25 ml of the
potassium dihydrogen
phosphate stock solution
and 5 ml of the magnesium
sulfate stock solution to
each liter of distilled
water to be used in the
preparation of dilution
water.
4.	Deliver enough working
solution to each dilution
water bottle so that after
sterilization the bottle
will contain 99 + 2 ml of
dilution water.
5.	Place caps on bottles
loosely.
6.	Sterilize in autoclave.
7.	Remove from autoclave
tighten bottle caps; cool
to room temperature.
8.	Store in cool place.
3a. A 10 ml or 5 ml pi pet is satisfactory for
delivery of both of these stock solutions
provided that it has graduation marks to
deliver the proper amount. Use separate
pipets for each solution to prevent contamination
of the stock solutions.
4a. Recommended dilution water bottles have a mark-
ing at the desired 99 ml quantity.
4b. Amount to be delivered to bottle before sterili-
zation cannot be stated exactly as evaporation
is different with differing conditions and auto-
claves. Ordinarily about 102 ml will be re-
quired.
6a. 15 minutes at 121°C
6b. Pressure reduced from autoclave gradually. This
is usually called "liquid cool" on autoclave dial
markings of automatic autoclaves.
8a. Dilution bottles ready for use. May be stored
indefinitely.
8b. Some evaporation losses may occur in time and in
these cases, sterile similarly prepared water
can be added. This is why a calibrated marked
bottle is desirable.

Page No. 9-15
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EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Membrane Filter Method
Page No. 9-16
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
12. Preparation of
M-FC medium.
1.
Prepare 0.2 normal
solution of sodium
hydroxide by adding 0.8
grams of solid sodium
hydroxide to 100 ml
distilled water.
la. Solution keeps indefinitely, should be protected
from evaporation losses with rubber stopper,
lb. CAUTION: sodium hydroxide is corrosive. Add
sodium hydroxide to the water, never water to the
sodium hydroxide,
lc. Unused solution may be stored until exhausted in
regrigerator and labeled as 0.2 N sodium
hydroxide.


2.
Prepare 1% Rosolic Acid
solution by dissolving 0.1
gram of Rosolic Acid
powder in 10 ml of 0.2
normal solution of sodium
hydroxide.
2a. Rosolic acid should be weighed on analytical
balance.
2b. Sodium hydroxide solution can be measured with
10 ml pipette.
2c. Unused Rosolic Acid solution can be kept up to
2 weeks if stoppered in refrigerator, and its
color remains a dark red; it is best prepared
freshly, however.
VIA.12.2

3.
Weigh 3.7 grams of Dehy-
drated M-FC Broth.
3a. Medium is hygroscopic (picks up moisture from
air) and should be stored in tightly stoppered
bottle, preferably in the dark, in a desiccator
(a closed jar or cabinet which contains materials
which take moisture out of the air).


4.
Place the weighed medium
in a clean, dry flask
having about 250 ml
capacity.
4a. This flask holds more than twice the volume of
the required solution because the medium expands
and foams when heated and space is required for
swirling of flask to mix contents.


5.
Add 1 ml of the 1%
solution of Rosolic Acid
to a 100 ml graduate, and
fill to the 100 ml mark
with distilled water.
5a. Note that this will be a final volume of 100 ml
and not 101 ml.
5b. This will be enough for 100 ml of culture medium,
or about 50 membrane filter plates. If different
amount of medium is required, adjust all
materials in proportion.
II.A.12.5
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EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Membrane Filter Method
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES

6.	Add a small amount of the
Rosoli c-Aci d-di sti11ed
water mixture to the flask
of weighed powder, and mix
until all the powdered
medium is dissolved from
walls of flask (no
sticking powder). Then
add the remainder of the
water and mix.
7.	Heat the medium with con-
stant agitation until the
boiling point is reached,
and then remove from heat
and cool promptly to below
45 C.
8.	Medium is ready for use.
7a. Agitation necessary to avoid burning the medium.
7b. Cooling is best by holding flask in a stream of
cool water.
Sa. Medium unused on day of preparation may be stored
up to 4 days if kept in refrigerator.
8b. Final pH of medium should be 7.4 j^O.l units.
This pH can be taken by utilizing a small portion
of the preparation and discarding after
measurement.

B. First-day procedures
1.	Equipment
Maintenance
2.	Assembly of
filtration
Material
1. Check, record, and adjust
incubator temperature, if
necessary.
1. Membrane filtration pro-
cedure equipment assembled
for analysis.
la. See A.l
la. Funnel clean and sterile
lb. Filtration flask and vacuum system operating
lc. Assembly of:
Data sheet, fecal coliform test
Sterile petri dishes
Sterile membrane filters with absorption pads.

Page No. 9-17
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EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Membrane Filter Method
Page No. 9-18
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
3. Sample collection
4. Preparation of
Laboratory Data
Sheet
1. Collect sample.
2. Record sampling
information.
3. Transport sample to
laboratory.
1. Fill in data sheet to
show sample information.
Sterile buffered distilled rinse water
Forceps and disinfectant container
Pencil, marking
Sample bottle
Sterile pi pets, 10 ml
Plastic bags
Burner, gas
Pan, pipet discard with disinfectant
la. Location selected by plant management,
lb. Sampling method as described in procedure
"Sample Collection and Handling for Bacterio-
logical Tests" and in the current edition of
Standard Methods.
2a. Most plants have standardized sample tags which
includes desired information, such as:
Collectors name
Date
Sample location
Time collected
Witness
Sample delivered to: 	
Time of delivery
2b. Tag may be retained as permanent record.
3a. Transported to laboratory without delay.
3b. Sample iced if delay of starting test is greater
than one hour.
3c. No longer than six hour delay from collection
time to laboratory delivery. A two hour addi-
tional time period is allowable from taking the
sample from the ice chest to completing membrane
filtration first-day procedures.
la. Information needed should be on sample tag.
lb. Minimal information on data sheet should include
-------
EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Membrane Filter Method
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
5.	Preparation of
laboratory bench
area
6.	Petri dish
preparation
2. Select sample volumes and
record on data sheet.
1. Disinfect laboratory bench
area.
1.	Set out required number of
sterile petri dishes on
laboratory bench.
2.	Place a sterile absorbant
pad in each petri dish.
'
source, date, collection time, name of sampler,
name of lab analyst, assigned sample number,
time of start of test, and sample volumes.
2a. According to fecal coliform density range
predicted for the sample.
2b. For fecal coliforms per 100 ml in the range:
Expected range per 100 ml
From To Sample Volume in Milliliters
2000 - 6000 1
400 - 1200 5*
200 - 600 10*
100 - 300 20*
40 - 120 50
20 - 60 100
Volumes showing asterisk (*) should be those
used as these will cover the range of counts
to demonstrate compliance or non-compliance with
effluent permit requirements.
la. Sponge, disinfectant solution, paper toweling.
la. Plastic dishes are purchased in a pre-sterilized
condition.
2a. Handle aseptically (not introducing bacterial
contamination) using a forceps which has been
stored in methanol or ethanol and flamed by
passing the forceps quickly through a flame,
keep approximately 1/2 of forceps in alcohol by
using jar or beaker.

Page No. 9-19
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EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Membrane Filter Method
Page No. 9-20
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES

3.
Using a sterile pi pet,
pipet approximately 2 ml
of M-FC medium over each
absorbant pad.
3a. Amount does not have to be precise as 2 ml is an
excess.
3b. Medium prepared and handled in accordance with
A.12.
3c. Keep plates covered.


4.
Gently tip uncovered
and allow any excess
medium to flow out of
plates.
4a. Undue tapping or shaking may spill too much
medium and leave plates without sufficient
medium.
4b. Cover plate. It is now ready for use.


5.
Label each dish with the
sample volumes to be
filtered.
5a. Use wax pencil or stick on label with pen.
5b. Label bottom (or base) of each plate.
Pad will not fall when plate is inverted.

7. Filtration
procedure
1.
Assemble filter assembly
upon filtration flask.
la. Sterile funnel units removed from wrapping,
lb. Using care to prevent contamination, such as
would be caused by fingers, touching of units to
equipment, etc.
lc. Unit should be connected to vacuum source and
have a means of vacuum disconnection, such as by
pinch clamp on the hose.


2.
Place membrane filter on
base of funnel apparatus.
2a. Funnel top removed carefully to avoid
contamination.
2b. MF should be grid or inked side up. MF handled
with flamed forceps and only by its outer 1/8
inch edge.
2c. Replace funnel top. Avoid over tightening


3.
Deliver measured volume of
a well shaken sample into
the funnel.
3a. Well shaken to insure even distribution of
bacteria.
3b. Poured gently into funnel, either by pipeting or
by use of a presterilized graduated cylinder (use
Kraft paper or foil hood). Avoiding splashing
of sample, and if graduate cylinder is used,
rinsed several times with small amounts of
sterile buffered distilled water which are also
poured into the funnel.
V.B.7.3
-------
EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Membrane Filter Method
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
8. Plating procedure
4.	Apply suction.
5.	Rinse funnel.
6.	Remove MF.
7.	Reassemble funnel.
1. Remove MF.
3c. If small sample portions are to be used (less
than 5 ml), a small amount of sterile buffered
distilled must be added to the funnel prior to
sample addition and then a gentle swirling of
the funnel to distribute the bacteria present.
4a. Vacuum applied only after sample has been com-
pletely delivered to funnel.
4b. Wait for complete evacuation of sample from
funnel.
5a. Three separate rinses with sterile buffered
distilled water. Complete evacuation of water
must occur between each application of rinse-
water using about 20 ml for each rinse.
5b. Vacuum supply shut off after last rinse.
6a. Handle gently with flamed forceps only on outer
1/8 inch of MF edge.
6b. Lifted gently from funnel base to break residual
vacuum before lifting.
7a. Unit is ready for next sample and sterilization
will not be required.
7b. If unit is not used within an hour it is
advisable to re-sterilize.
7c. Handling of funnel top is critical in that no
contamination should occur. Avoid handling in
funnel surfaces that receive sample and do not
lay on table that may have residual germicide.
A ringstand with split ring or resting on the
funnel top only on its base after hand lifting
are recommended methods.
la. This was done as part of the filtration procedure
(B.7.6) and held with one hand as the funnel is
reassembled (8.7.7).
V.B.7.7
Page No. 9-21
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EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Membrane Filter Method
Page No. 9-22
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
9. Incubation
procedure
2.	Remove cover from petri
dish with prepared M-FC
medium.
3.	Plate MF over medium.
1. Place plate/plates in pro-
tective plastic bag and
incubate at 44.5 C.
2a. With practice this can be done with one hand
while the MF is held in the other. As an aid for
easier operations, the plates can be organized in
the order of plating and their tops can be
loosened.
3a. Allow the side of the MF opposite the forceps to
touch the saturated absorption pad, and with a
rolling action, plate the entire membrane onto
the pad. The grid (inked squares) must be up.
3b. Rolling action will tend to expel air bubbles
from under the MF and allow an intimate contact
between medium and MF. Air pockets must be
removed for recovery of all fecal coliform
organisms.
3b. Air pockets removed from underside of MF by
gently lifting some or all of MF and re-rolling
upon medium. At no time must a forceps be used
to "smoothout" the surface of the MF. Any move-
ment on the delicate surface of the MF by the
forceps will result in a line growth of bacteria
instead of the easily countable discrete colony
form.
la. Completed plates must be incubated within 30
minutes from the time they are filtered.
lb. A variety of bags and bag closure devices exist.
One of the simplest is a "Whirl-Pac" which has
metal wire fold covers to keep rolled end sealed
and leakproof.
lc. Bags containing plates must be totally immersed
within the incubator's water. It is usually
necessary to arrange some weight, such as a test
tube rack, to keep the plates under water.
Id. All petri dishes must be incubated in the in-
verted position (pad, medium, and MF) now on top
of the plate) so that droplets cannot fall on
surface of MF.

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EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Membrane Filter Method
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES


16. Observe immersed bag for a short time to observe
that a constant bubbling action is not occuring
to indicate bag leakage. Reseal if this is
occurring and recheck.
If. Allow to incubate for 24 hours +2.0 hours.

C. Second Day Procedure
1. Counting procedure
1.	Remove bag from water bath
incubator.
2.	Remove plates from plastic
bag.
3.	Select plates which have
from 20 to 60 colonies.
4.	Count fecal coliform
colonies with microscopic
aid.
la. Be sure the incubation has been within the limits
of 24 hours + 2 hours.
lb. Handle carefully to avoid droplet splattering
within plates.
2a. Set plates on table so that colonies (growth)
are visible.
3a. This ability comes with experience, but plates
which are overcrowded or the ones which have
fewer colonies should be readily apparent.
It is necessary only to record plate counts
within these ranges. If this is not possible
other counts can be used as described further.
4a. Binocular wide field dissecting microscope pre-
ferred.
4b. Use a 10-15 X magnification with fluorescent
1ighting.
4c. Scan membrane with a back-and-forth movement
over the grids, line by line, so as to cover
the membrane completely without missing any area.
4d. All blue or blue-tinted (blue-green, purple, etc.)
colonies are counted as fecal coliforms. Color-
ation may be deep or light and can be all over
or partially cover the colony. Some can even
have the coloring appear in flecks on the
surface.
V.C.I.4
Page No. 9-23
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EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Membrane Filter Method
Page No. 9-24
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
7F:AI.\;ING
guide :;ctes

5- Select colony count/counts
to use. Use formula to
calculate count per 100 ml.
5a. Formula
Fecal Coliforms per 100 ml =
100 x colony count
number of milliliters (mis) filtered
5b. Select colony count which falls within the 20-60
range:
Example
mis Filtered No. Colonies
20 TNTC (indicates
too numerous
to count)
10 TNTC
5 35
Use: 35 colonies with a 5 ml sample volume
Fecal coliform = 100 x 35 = 700
per 100 ml 5
5c. If more than one plate has colony numbers within
the range, add the results.
Example
mis Filtered No. Colonies
20 45
10 23
5 9
20 ml + 10 ml =30 ml
45 colonies + 23 colonies = 68
Fecal coliforms per 100 ml = 100 x 68 = 227
30
Use: 230 (nearest two significant figures)

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EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Membrane Filter Method
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
5d. If no counts were obtained within these ranges,
the following procedure should be followed:
TRAINING
GUIDE NOTES
All above 60 Colonies
Use that count which is closer to the maximum
60 count.
Example
20 ml	TNTC
10 ml	150
5 ml	72
Use: 72 colonies with a 5 ml sample volume
fecal coliforms/100 ml = 100 x 72 = 1440
"T
or 1400 fecal coliform per 100 ml
All below 20 Colonies
Use that count which is closer to the 20 count.
Example
20 ml	15
10 ml	8
5 ml	0
Use: 15 colonies with a 20 ml sample volume
which gives 75/100 ml fecal coliforms.
All Plates with a Zero Count of Fecal Coliform
Assume that the largest volume delivered has one
colony. Use this in calculations and call the
result <(les$ than). If all three plates show
a zero count the fecal coliform count would be
< 5 (calculation: 100 x^l).
Page No. 9-25
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EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Membrane Filter Method
Page No. 9-26
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
6. Record colony counts on
data sheet.
£ There is no such thing as a "standard" data
sheet for bacteriological tests. A simplified
data sheet is shown below:
Sample Type
Station
Fecal Coliform Test
Membrane Filter (MF) Procedure
Lab. No.
Collection Date
Received_
pH	
_Description_
Time
APM. Temp.
APM. Examined
APM.
Observations
ml s
Filtered
Colony Count
Remarks





















Results:
Fecal Coliform (MF)
-------
EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Membrane Filter Method
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES

7. Record fecal coliform
count/100 ml.
7a. Record on designated data sheet for your agency.
7b. Record to nearest two significant figures.
Examples
266.6 will be 270
20.09 will be 20
299.4 will be 300

Page No. 9-27
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TRAINING GUIDE
SECTION	TOPIC
I	Introduction
11*	Educational Concepts - Mathematics
HI	Educational Concepts - Science
IV	Educational Concepts - Communications
V*	Field and Laboratory Equipment
VI*	Field and Laboratory Reagents
Vii	Field and Laboratory Analysis
VIII	Safety
IX	Records and Reports
*Training guide materials are presented here under the headings marked*.
These standardized headings are used throughout this series of procedures.
Page No. 9-28
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FFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Membrane Filter Method
Educational Concepts - Mathematics	Section jj

TRAINING GUIDE NOTE
REFERENCES/RESOURCES
A.12.5
Since 3.7 grams of MFC powdered medium and 1 ml of
\% Rosolic Acid is required to prepare 100 ml of
MFC broth, it is possible to calculate weights and
volumes to prepare any requirement based upon the
number of plates desired. Calculations are based
on knowing the above figures and the requirement
of 2.0 ml of broth for each plate.
For rapid calculations the following two formulas
can be used:
(1)	No. of plates desired X 0.074 = Grams MFC
(2)	No. of plates desired X 0.02 = ML Rosolic Acid
EXAMPLE
If 125 plates of MFC are required:
125 X 0.074 = 9.25
=9.3 grams MFC medium required
125 X 0.02 = 2.5 mis 1% Rosolic Acid
required
Note: Due to the difficulties involved in weighing
very small portions as, for Instance, .074
grams of MFC for one plate requirement, 1t
would be wise to prepare at least 10 plates
(.7 gr. MFC and 0.2 ml Rosolic Acid) as a
minimum requirement.

Page No. 9-29
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EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Membrane Filter Method
Field and Laboratory Equipment	Section y

TRAINING GUIDE NOTE
REFERENCES/RESOURCES
A.1.1
A.1.2
Incubator should be kept out of drafts or direct
sunlight in order to prevent temperature inside the
incubator from changing outside the temperature
range specified (44.5 C + 0.2 C).
An accurate solid heat sink may be used. This is
constructed of a solid aluminum block and does not
contain water for transference of heat. Plastic
bags, for this reason, are no longer required when
using this type of incubator. Since there are no
provisions for a high humidity chamber in this type
of incubator, it is important to use only the types
of petri dishes having a tight attachment of cover-
to-base thus preventing loss of moisture during the
incubation.
Standard Methods for the
Examination of Water and
Wastewater 14th Ed. (1975)
APHA, WPCF, AWWA, p.880 ff
(Hereafter refferred to as
Std. Meth. 14: (Page No. )
Std. Meth. 14: 937
A.1.3
Mercury bulb thermometer usually used in most
incubators and a recording thermometer is
acceptable. Thermometers must be calibrated against
a mercury bulb thermometer which is (or calibrated
against) a National Bureau of Standards issued and
used with the certificate and correction chart.

A. 3
Autoclaves differ greatly in design and in method
of operation. Some are almost like home-style
pressure coolers; others are almost fully auto-
matic.
Std. Meth. 14: 881
A.4
Distilled water must not contain substances pre-
venting bacterial growth or be highly nutritive.
There are required procedures to testing distilled
water and should be undertaken only by professional
bacteriologists or in laboratories where this is
done regularly.
Std.Meth. 14: 888
A.6.2
Glassware can be checked for bacteriostatic or
inhibitory residues by a bacteriological test
procedure which, like the distilled water suita-
bility test, should be undertaken only by pro-
fessional bacteriologists or in laboratories where
this test is done on a regular basis.
Std. Meth. 14: 885
Page No. 9-30
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EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Membrane Filter Method
Field and Laboratory Equipment	Section v

TRAINING GUIDE NOTE
REFERENCES/RESOURCES
A.9.1-6
Wide-mouth, glass-stoppered bottles suggested, but
other styles accepted. Bottle must be heat stable
to sterilizing conditions and not be toxic or
nutritive to organisms natural to the sample.
If glass-stoppered bottles are used, a strip of
paper should be placed in the neck of the bottle
before placing the stopper in place in preparation
for sterilization. This prevents the glass stopper
from "freezing" in place during sterilization.
The paper strip is descarded at the time of sample
collection.
Std. Meth. 14: 884
14: 904
A.10.1-5
This procedure is described in terms of reusable
glass pipets. However, single-service, pre-
packaged, glass or plastic pipets may be used.
In the case of single-service pipets, they will be
sterile when purchased, are used one time, and
discarded after use. Accordingly, in the step-by-
step procedures, disregard any instructions about
pi pet preparation if these pipets are used.

Paqe No. 9-31
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EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Membrane Filter Method
Field and Laboratory Equipment
Section V
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
B.7.7
If graduate is labeled TC (To Contain), follow the
rinsing procedure. If it is labeled TD (To Deliver)
slowly add the complete sample, gently tap off the
last drop and do not rinse.
When the filter holding unit is disassembled after
sample filtration, the worker's hands must be free
to manipulate the membrane filter. Upon disassembly
of the filter holding unit, many workers place the
funnel element, inverted, on the laboratory bench.
Some workers, to prevent bacterial contamination,
prefer a rack or a support to keep the funnel element
from any possible source of contamination. A split
ring on a ring stand is a convenient rack for this
purpose.
The dashed circle indicates the effective filtering
area. The dashed back-and-forth line indicates the
colony counting pathway.
SUCTION FLASK RING STAND WITH SPLIT RING
Paqe No. 9-32
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EFFLUENT MONITORING PROCEDURE: Fecal Coliform Test by the Membrane Filter Method
Field and Laboratory Reagents	section

TRAINING GUIDE NOTE
REFERENCES/RESOURCES
A.12.2
Rosolic acid may be omitted from the medium if
minimal background colony counts occur and
equivalent results are obtained without it.
Std. Meth. 14: 894
Paae No. 9-33
-------
A PROTOTYPE FOR DEVELOPMENT OF
ROUTINE OPERATIONAL PROCEDURES
for the
CALCULATION OF THE GEOMETRIC MEAN
OF COLI FORM COUNTS
by the
USE OF LOGARITHMS
as applied In
WASTEWATER TREATMENT FACILITIES
and in the
MONITORING OF EFFLUENT WASTEWATERS
Developed by the
National Training and Operational Technology Center
Municipal Operations and Training Division
Office of Water Program Operations
U.S. Environmental Protection Agency
ST.EMP.1.3.77	Page No. 10-1
-------
EFFLUENT MONITORING PROCEDURE: Calculation of the Geometric Mean of
Coliform Counts by the Use of Logarithms
This procedure was developed by:
NAME	Joseph F. Santner
ADDRESS EPA, UWPU, NTOTC, Cincinnati, Ohio 45268
POSITION Mathematical Statistician
EDUCATION & TECHNICAL BACKGROUND
BS - St. Louis University
MS - St. Louis University
4 years - U.S. Army
3 years - Operations Research Analyst
7 years - University Professor - Mathematics
12 years - Statistical consultant and training of Federal,
State, and Local personnel in the principles and
practice of statistical analysis
Page No. 10-3
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EFFLUENT MONITORING PROCEDURE : Calculation of the Geometric Mean of
Coliform Counts by the Use of Logarithms
1.	The object of this procedure is the calculation of the geometric mean of
fecal coliform counts when using logarithms.
2.	Brief description of the procedure.
How to use logarithms (or logs) and find the geometric mean (or GM) of
n fecal coliform counts, where each count is greater than or equal to one.
GM (of Ni, N2,. . Nn) = anti-logH°9 N1 + 1o9 N? + • • • + 1o9 N
In order to complete the calculations on the right hand side of the
equation, four operations are necessary.
A.	Determine the log for each of the n fecal coliform counts.
B.	Add or sum the n logs
C.	Divide the sum by sample size equal to n
D.	Find the anti-log of the answer to step C.
Let the first fecal coliform count = N-j
Let the second fecal coliform count = N2
etc.
Let the last fecal coliform count = Nn
Let n equal the total number of such fecal coliform counts or
n = sample size. The formula for the GM when using logs is:
Page No. 10-4
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EFFLUENT MONITORING PROCEDURE: Calculation of the Geometric Mean of
"	Coliform Counts by the Use of Logarithms
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
A. Finding the log of a
fecal coliform count
1. Determine d = number of
digits to the left of the
decimal point
2. Calculate C = d-1 (C is
called the characteristic
by mathematicians)
Examples
la.	if N] =23 then d = 2
lb.	if N2 = 122 then d = 3
lc.	if N3 = 17,100 then d = 5
2a.	if N] = 23 then d = 2 and C = 2-1 =
2b.	if Ni = 122 then d = 3 and C = 2
2c.	if Nt = 17,100 then d = 5 and C = 4
Page No. 10-b
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EFFLUENT MONITORING PROCEDURE: Calculation of the Geometric Mean of
Coliform Counts by the Use of Logarithms
Page No. 10-6
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING
GOALS/SPECIFICATIONS

TRAILING
GUIDE NOTES
A. (Cont'd.)
3. Locate N in the table margia
The first two digits in the
first column in the left
margin and the third digit
in the first row. Note that
trailing zero's can be added
or deleted in order to have
the necessary three digits
for entry into the tables.
3a. For N] =23, there
are only 2 digits
so we must add a
trailing zero. Lo-
cate 23 cr the
first two digits
in the left margin
and the third or
last digit 0, in
the top row.
3b. For N2 = 122, the
first two digits
or 12 are located
in the left margin
and the third digit
or 2 is located in
the top row.
3c. For = 17,100 the
first two digits or
17 are located in
the left margin.
The third digit is
a 1 and is located
in the first row.
All digits after
the third are de-
leted or these
trailing zeros are
ignored.
N.
1.(77] Q]
10
00 COO
00 432
GO S60
01 234
Ng->-|i2l
04
139
04 532
04
0V2
05 303
07
91S
03 279
03 KVj
03 991
11
294
11 727
12 057
12 335
14
14 C13
14 922
15
229
15 53 i
15
17
609
17 S-.93
18
1S4
IS 469
16
20
412
20 es3
20 952
21 219

23 015
23 300
23
553
23 805
0 18
25
527
25 76S
20
007
26 245
19
27
875
23 103
23
330
23 556
20
30
103
30 320
30
535
30 75o
21
32
222
32 42S
32
634
32 S33
22
34
242
34 439
31
635
34 S30
N-|-41D
36
173
36 361
36
54J
36 736
24
3S 021
33 202
33
3-2
33 fOl
25
3w
794
39 967
40
110
40 312
26
41
497
41 664
41
830
41 i}%
27
43
136
43 297
43
457
43 CIO
2S
44
716
44 871
45
025
45 179
29
46
210
46 339
40
0
46 637
30
47
712
47 857
4S
001
4S 144
31
49
136
49 276
49
415
49 551
32
50
515
50 f,51
50
7m'i
50
33
51
hoi
51 9 S3
52 114
52 244
34
53
149
53 275
53 403
53 529
35
54
407
54 531
54
('=>1
54.777
36
55
630
55 751
55
371
55 991
37
56 820
56 937
57
054
57 17?
38
57
978
53 092
53
206
58 329
39
59
106
59 218
59
329
59 439
40
GO 206
GO 314
CO
423
60 531
41
61
27S
61 334
61
490
61 595
42
62 325
02 42S
62
531
62 634
43
63 317
63 443
63
513
63 6U
-------
EFFLUEI1T MONITORING PROCEDURE: Calculation of the Geometric Mean of
Coliform Counts by the Use of Logarithms
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
rp.AiNir;G
^ N1 ^ N3
lGH CD
A. (Cont'd.)
4. Read the 5 digit number
within the body of the
table at the intersection
of the row and column
determined in step 3. We .
label this number.M. The
mathematicians call it the
mantissa.
4a. For N] = 23 then
M = 36173
4b. For N£ = 122 then
M = 08636
4c. For N3 = 17,100
then M = 23300
N^
N.
10
&
13
14
15
16
~iia
18
19
20
21
22
^-123
24
25
26
27
28
20
31
32
33
3t
35
3G
3'
3S
39
40
41
42
43
STn2
m
00 000 CO 432 00 S60 01 2*1
04 139 04 532
07 918 05 279
11 394 11 727 12 057
05 3DS
OS 9'Jl
12 3S5
14 613 14 922 15 220 15 534
17 609 17 SOS IS 184 IS 4f.9
20 412 20 683 20 952 21 219

23 045
25 527 25 763
27 875 28 103
23 553 23 805
26 007 26 245
28 330 23
30 103 30 320 30 535 30 750
32 222	32 42$	32 634	32 838
34 2-12	31 439	34 (,35	31 830
36 1/3^	36 301	3(1 519	36 736
38	021	38 202	38 3S2	3S 501
39	794	39 967	40 140	40 312
41 497	41 664	41 830	41 996
43	136	43 297	43 457	43 616
44	716	44 871	45 025	45 179
46 210	46 3S9	40 538	46 Gb7
47 712 47 857 4S 001 4S 144
49	136	49 270	-19 415	49 554
50	515	59 651	50 786	f.0 920
51	851	51 983	52 114	52 244
53	148	53 275	53 403	53 529
54	407	54 531	54 654	S4.777
55	630	55 751	55 871	55 991
56	S20	56 937	57 054	57 171
57	978	AS 092	58 206	58 320
59 106	59 218	59 329	59 439
CO 206 60 314 CO 423 CO 53!
61	278 61 3S1 61 WO 6! 595
62	325 62 428 02 531 62 634
63	347 63 448 63 545 63 619
Page No. 10-7
-------
EFFLUENT MONITORING PROCEDURE: Calculation of the Geometric Mean of
Coliform Counts by the Use of Logarithms
Page No. 10-8
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
A. (Cont'd.)
5. Log N = C.M
5a. Log 23 = 1.36173
5b. Log 122 = 2.08636
5c. Log 17,100 = 4.23300
t \
C obtained M obtained
in step 2 in step 4

B. Sum the n logs
1. It is assumed that addition
is known.


C. Divide the sum of
the logs by n,
sample size
1. It is assumed that division
is known


D. Finding the anti-log
of a positive number
1. Determine M = the number to
the right of the decimal
point.
la. If we want the anti-log of 3.11394 then
M = 11394
lb. If we want the anti-log of 2.32428 then
M = 32428
1c. If we want the anti-log of 2.56036 then
M = 56036

-------
EFFLUENT MONITORING PROCEDURE: Calculation of the Geometric Mean of
~~ "	Coliform Counts by the Use of Logarithms
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
10 00 000 00 432 00 SCO 01 2M 01 703
01 139 04 532 Ot 922
07 018 OS 27'J OS (Be,
in am u 727 12 001
0,-> 3'IS 0.) O'.K)
Ob y.u p.';ii2
12 3s5 12 710
14 013 14 922
17 C09 17 898
20 412 20 GS3
l.» 229
15 531 )¦> xw,
18 400 IS
21 219 21 1M
18 184
20 952
23 015 23 300
21 ,'.27 25 76S
103
23 553
21; 00:
28 330
23 805 21 055
2i i 215 21'. I >'2
2S 5:-<> 28 7SO
30 330
(0 !«;•(
30 535
32 G34 P.2 8!iS 33 0 i I
31 035 31 830 35 : 41; 8H0
12 47
4s (ill 48 H i 48 28.
49 551 49 c.
(Nearest ^alue
is 55991 )\
3a. If M = 11394 then
N = rc = 130
since r = 13 and
c = 0
3b. If M - 32428 then
N = 211 since r =
and c = 1
3c. If M = 56036 then
N = 363 since r = 36
and c = 3
4a. For the anti-log of
3.11394 then C = 3
4b. For the anti-log of
2.32428 then C = 2
4c. For the anti-log of
2.56036 then C = 2
5a. If C = 3, then d = 4
5b. If C = 2, then d * 3
5c. If C = 2, then d = 3
6a. Anti-log 3.11394 =
1300. The 130 was
determined in step 3a
and the decimal point
was placed so that 4
digits (see 5a) are to
the left of it. In this
case a trailing zero was added
Page No. 10-9
-------
EFFLUENT MONITORING PROCEDURE: Calculation of the Geometric Mean of
Coliform Counts by the Use of Logarithms
Page No. 10-10
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
D. (Cont'd.)
6. (Cont'd.)
6b. Anti-log 2.32428 = 211. Step 3b followed by
step Eb.
6c. Anti-log 2.56036 = 363. By combining steps
3c through 5c.

-------
—Common logarithms of numbers—
N.
10
11
12
13
14
15
16
17
IS
19
XO
21
22
23
2i
25
26
27
28
29
30
31
32
33
34
35
36
37
38
36
40
41
42
43
44
45
46
47
48
49
M
L. 0
00 000 00 432 00 860 01 284 01 703
02 110 02 531 02 938 03 342 03 743
04 139
07 918
11 394
14 613
17 609
20 412
23 045
25 527
27 875
04 532
08 279
11 727
14 922
17 898
20 083
23 300
25 768
28 103
04 922
08 636
12 057
15 229
18 184
20 952
23 553
26 007
28 330
05 308
08 991
12 385
15 534
18 409
21 219
23 805
26 245
28 556
05 690
09 342
12 710
15 836
18 752
21 484
24 056
26 482
28 780
06 070
09	691
13 033
10	137
19 033
21 748
24 304
26 717
29 003
06 448
10 037
13 354
16 435
19 312
22 Oil
24 651
26 951
29 226
06 819
10 380
13 672
16 732
19 590
22 272
24 797
27 184
29 447
07 188
10 721
13 988
17 026
10 866
22 531
25 042
27 416
29 667
07 555
11 059
14 301
17 319
20 140
22 789
25 285
27 646
29 885
30 103 30 320 30 535 30 750 30 9C3
31 175 31 387 31 597 31 806 32 015
32 222
34 242
36 173
38	021
39	794
41 497
43	136
44	710
46 240
32 428
34 439
30 361
38	202
39	9
-------
—Common logarithms of numbers-
N.
L. 0
60
61
52
53
64
66
66
67
68
69
60 897 69 984 70 070 70 157 70 243
70 329 70 415 70 501 70 586 70 672
70	757	70 842
71	600	71 G84
72	428	72 509
73	239	73 320
74	036	74 115
74	819	74 896
75	587	76 664
76	343	76 418
77	085	77 159
70	927
71	767
72	591
73	400
74	194
74 974
76 740
76	492
77	232
71 012
71	850
72	673
71 096
71	933
72	754
73 480	73 560
71 273	74 351
76	051	75 128
75 815	75 891
70 567	76 641
77	305	77 379
71,181	71 265
72 016	72 099
72	835	72 916
73	640	73 719
74	429	74 507
75	205	75 282
75	967	76 042
76	716	76 790
77	452	77 525
71	349
72	181
72	997
73	799
74	586
75	358
76	118
76	864
77	697
71	433	71 517
72	263	72 346
73	078	73 159
73	878	73 957
74	663	74 741
75	435	75 511
76	193	76 268
76	938	77 012
77	670	77 743
77 815 77 887 77 960 78 032 78 104
78 176 78 247 78 319 78 390 78 462
78	533	78 604
79	239	79 309
79	934	80 003
80	618	80 686
81	291	81 358
81	954	82 020
82	607	82 672
83	251	83 315
83 885	83 948
78	675
79	379
80	072
80	754
81	425
82	086
82	737
83	378
84	Oil
78	746	78 817
79	449	79 518
80	140	80 209
80	821	80 889
81	491	81 558
82	151	82 217
82	802	82 866
83	442	83 506
84	073	84 136
78	888	78 958
79	688	79 657
80	277	80 346
80	956	81 023
81	624	81 690
82	282	82 347
82	930	82 995
83	569	83 632
84	198	84 261
79 029
79	727
80	414
81	090
81	757
82	413
83	059
83	696
84	323
79 099	79 169
79	796	79 865
80	482 80 650
81	158	81 224
81	823	81 889
82	478	82 543
83	123	83 187
83	769	83 822
84	386	84 448
84 510 84 572 84 634 84 696 84 757
84 819 84 880 84 942 85 003 85 065
85 126 85 187	85 248	85 309	85 370
85	733 85 794	85 854	85 914	85 974
86	332 86 392	86 451	86 510	86 570
86	923	86 982	87 040	87 099	87 157
87	506	87 564	87 622	87 679	87 737
88	081	88 138	88 195	88 252	88 309
88	649	88 705	88 762	88 818	88 874
89	209	89 265	89 321	89 376	89 432
89 763	89 818	89 873	89 927	89 982
85	431	85 491
86	034	86 094
86	629	86 688
87	216	87 274
87	795	87 852
88	366	88 423
88	930	88 986
89	487	89 542
90	037	90 091
85	552
86	153
86	747
87	332
87	010
88	480
89	042
89	597
90	146
85	612	85 673
86	213	86 273
86	806	86 864
87	390	87 448
87	967	88 024
88	536	88 £93
89	008	89 154
89	653	89 708
90	200	90 255
90 309 90 303 90 417 90 472 90 526
90 580 90 634 90 687 90 741 90 795
90	849	90 902	90 956	91 009	91 062
91	381	91 434	91 487	91 540	91 593
91	-SOS 91 960	92 012	92 065	92 117
92	428	92 480	92 531	92 583	92 634
92 942	92 993	93 044	93 095	93 146
S3 450	93 500	93 551	83 601	83 651
S3 952	94 002	94 052	94 101	94 151
94 448 94 498	94 547	94 596	94 645
04 909	94 988	95 036	95 085	95 134
91 116	91 169
91	645	91 698
92	169	92 221
92	686	92 737
93	197	93 247
93 702	93 752
91 222
91	751
92	273
92	788
93	298
93 802
91 275	91 328
91	803	91 855
92	324	92 376
92	840	92 891
93	349	93 399
S3 852	93 902
94 201
94	694
95	182
94 250 94 300
94	743 94 792
95	231 95 279
94 349
94	841
95	328
94 399
94	890
95	376
96 424 95 472 95 621 95 569 95 617
95 665 95 713 95 761 05 809 95 856
85 904	95 952
96 379	96 426
96	848	96 895
97	313	97 3.r)9
97	772	97 818
98	227	98 272
98	077	98 722
99	123	99 i
-------
EFFLUENT MONITORING PROCEDURE: Calculation of the Geometric Mean of
Coliform Counts by the Use of Logarithms
An example of the calculations for operating procedure A, B, C, and D follows.
GM (23, 122, 17100) = Anti-logjTog 23 + log 122 + log 17100~J
(see A5a, 5b, 5c)
GM (23, 122, 17100) = Anti-loqj~L36173 + 2.08636 + 4.23300~j
(See procedure B and C)
GM (23, 122, 17100) « Anti-log 2.56036
(See D6c)
GM (23, 122, 17100) = 363.
The following practice problems should be solved to make sure that the program
of action is mastered.
1)	GM (1, 4) = 2
2)	GM (1, 10, 100) = 10
3)	GM (10, 10, 10) = 10
Some checks for gross errors.
1)	GM lies between the largest and smallest value. For the problem
GM (23, 122, 17100) = 363 the largest = 17,100 and the smallest = 23.
Since 363 lies between these two, there is no gross error.
2)	GM is less than the arithmatic mean* (AM). AM = 23 + 122 + 17100 = 5748.3
3
GM = 363 is less than AM * 5748.3. Hence, there is no gross error.
*GMsAM if all conform counts are equal as illustrated in practice problem
number 3.
Paae No. 10-
-------
A PROTOTYPE FOR DEVELOPMENT OF
ROUTINE OPERATIONAL PROCEDURES
for the
MEASUREMENT OF FLOW IN AN OPEN CHANNEL BY
PARSHALL FLUME
as applied 1n
WASTEWATER TREATMENT FACILITIES
and 1n the
MONITORING OF EFFLUENT WASTEWATERS
Developed by the
National Training and Operational Technology Center
Municipal Operations and Training Division
Office of Water Program Operations
U.S. Environmental Protection Aqencv
EN.FM.EMP.2a.8.76	Page No. 11-1
-------
EFFLUENT MONITORING PROCEDURE: Flow Measurement in an Open Channel by
Parshall Flume
This Procedure was developed by:
NAME	Charles E. Sponagle
ADDRESS EPA, OWPO, NTOTC, Cincinnati, Ohio 45268
POSITION Sanitary Engineer-Instructor
EDUCATION AND TECHNICAL BACKGROUND
B.C.E - Manhattan College, 1943
M.S. in C.E. - University of Minnesota, 1948
Professional Registration: State of New York
With Federal Water Pollution Control Program since 1948, with
various assignments at Program Headquarters, Regional Offices,
and Field Stations, including positions as
Staff Engineer, then Chief, Water Quality Section
Denver Regional Office
Staff Engineer, then Regional Construction Grants Program
Director, Denver Regional Office
Regional Construction Grants Program Director,
Cincinnati Regional Office
Director, Colorado River Basin Water Quality Control Project,
Denver, Colorado
Industrial Wastes Consultant, Technical Advisory and
Investigations Branch, Cincinnati, Ohio
Participation in and Direction of numberous in-plant industrial
waste surveys and stream studies 1n New York, Colorado, New
Mexico, Maine, Utah
With National Training & Operational Technology Center,
September 1969 to date.
Page No. 11-3
-------
EFFLUENT MONITORING PROCEDURE: Flow Measurement in an Open Channel by
Parshall Flume
1.	Objective: To enable the student to obtain the flow rate in an open
channel by means of a pre-installed Parshall Flume.
2.	Description of Procedure:
The depth of liquid is measured at a stipulated point (or points) within the
Flume. This measurement is then used to obtain the rate of flow in the
channel.
a.	This Procedure deals specifically with 6-inch through 8-foot flumes,
since practically all wastewater treatment plant influent and effluent
flows can be measured by flumes of this size. Operating principles of
larger and smaller size flumes are exactly the same. For these latter
however, some differences in procedures are involved, consisting of a
change of location for measurement of the downstream head, and use of
different discharge tables.
b.	Flows obtained by visual observation of liquid depth are considered
herein. Use of devices which automatically provide a continuous record
of either head or flow is not included.
General Description of Equipment used in the Procedure:
1)	Parshall Flume.
2)	Means for visually observing depth of flow, such as a staff gage
or a float gage.
Page No. 11-4
-------
EFFLUENT MONITORING PROCEDURE: How Measurement in an Open Channel by Parshall Flume
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
A. Basic Elements



1. Units of Flow


II.A.1
Measurement


(p. ID
2. Description of



Process


I.A.2



(p. 10)
3. Structure of



Flume


V.A.3



(p. 24)
4. Terminology ami


Definitions


V.A.4



(p. 25)
5. Operating



Principles


V.A.5



(p. 26)
6. Staff Gage


V.A.6



(P- 27)
7. Float Gage


V.A.7



(p. 27)
B. Preparation for
Measurement
1. Physical
Conditions
1.	Observe flow upstream of
flume
2.	Remove any objects causing
disturbance of flow
3.	Inspect flume for deposits
of solids
la. Reasonably smooth or streamline flow,
lb. Flow distributed reasonably uniformly across
channel
3a. No build-up of sediment in structure
III.B.1.1
(p. 22)
[II.B.1.3
(p. 22)
Page No. 11-5
-------
EFFLUENT MONITORING PROCEDURE: Flow Measurement in an Open Channel by Parshall Flume
Paqe No. 11-6
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES

4.	Determine flow condition
5.	Inspect stilling well, clean
if necessary
4a. Free flow or submerged flow.
5a. Connection to channel not clogged.
5b. No deposits
5c. No objects interfering with float
III.B.1.4
(p. 22)
III.B.1.5
(p. 23)
C. Flow Measurement -
Free-Flow Condition,
Using Staff Sage
1.	Determination of
upstream Head, Ha
2.	Determination of
flow rate.
1.	Read gage division at which
liquid surface intersects
gage.
2.	Calculate Ha
1. Use appropriate table.
la. To nearest division.
2a. From staff gage reading,
la. In unit desired.
II.CJ ,
(p- 12)
II.C.2
(p- 15)
D. Flow Measurement -
Free-Flow Condition,
Using Float Gage
1.	Determination of
upstream head, Ha
2.	Determination of
flow rate
1.	Read tape division opposite
index on float gage
2.	Calculate Ha
1. Use appropriate table
la. To nearest division.
2a. From float gage reading
la. In unit desired
II.D.l
(p. 16)
II.C.2
(P• 15)




-------
EFFLUENT MONITORING PROCEDURE: piow Measurement in an Open Channel by ParshalI Flume
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
E. Flow Measurement -
Submerged-Flow Con-
dition, using Staff
Gages




1. Determination of
upstream head, Ha
1.
Read gage division at which
liquid surface intersects
gage.
la. To nearest division,
lb. At the same time as for Hb
II.C.l
(p. 12)

2.
Calculate Ha
2a. From staff gage reading.

2. Determination of
downstream head,
«b
1.
Read gage division at which
liquid surface intersects
gage.
la. To nearest division,
lb. At the same time as for Ha
II.C.l
(p. 12)

2.
Calculate H5
2a. From staff gage reading.

3. Determination of
flow rate.
1.
Calculate percent sub-
mergence
la. Percent submergence = Hk x 100
Ha


2.
Consult appropriate chart
or table.

II.E.3.2
(p. 17)

3.
Read submerged flow value,
or obtain correction to be
applied to free-flow value.



4.
When a correction factor is
obtained, use Ha and find
free-flow from Table 1.



5.
Multiply this free-flow
value by the correction
factor to obtain the sub-
merged flow.






Page No. 11-7
-------
EFFLUENT MONITORING PROCEDURE: p]ow Measurement in an Open Channel by Parshall Flume
Page No. 11-8
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
F. Flow Measurement -
Submerged-Flow
Condition, using
Float Gages



1. Determination of
upstream head, Ha
1.	Read tape division opposite
index on gage.
2.	Calculate Ha
la. To nearest division,
lb. At the same time as for %
2a. From float gage reading.
II.D.1
(p. 16)
2. Determination of
downstream head,
»b
1. Read tape division opposite
index on gage.
la. To nearest division
lb. At the same time as for Ha
II.D.1
(p. 16)
2. Calculate
2a. From float gage reading

3. Determination of
flow rate.
1.	Calculate percent sub-
mergence.
2.	Consult appropriate chart
or table.
3.	Read submerged flow value
directly or obtain correction
to be applied to free-flow
value.
4.	When a correction factor is
obtained, use Ha and find
free-flow from Table 1.
5.	Multiply this free-flow value
by the correction factor to
obtain the submerged flow.
la. Percent submergence = Hb x 100
Ha
II.E.3.2
(p. 17)
-------
EFFLUENT MONITORING PROCEDURE: Flow Measurement In an Open Channel by ParshalT Flume
TRAINING GUIDE
SECTION	TOPIC
I*	Introduction
II*	Educational Concepts - Mathematics
III*	Educational Concepts - Science
IV	Educational Concepts - Communications
V*	Field & Laboratory Equipment
VI	Field & Laboratory Reagents
VII	Field & Laboratory Analysis
VIII	Safety
IX	Records & Reports
~Training guide materials are presented here under the headings marked*.
These standardized headings are used throughout this series of procedures
Paqe No. 11-9
-------
EFFLUENT MONITORING_PROCEDURE. Measurement of Flow in an Open Channel by Parshall Flume
INTRODUCTION		Section I
TRA1 FUNG GUIDE NOTE
REFERENCES/RESOURCES
A.2
Flow of a liquid in an open channel can be
measured in many cases by means of a specially-
shaped section known as a Parshall Flume. The flume
can be constructed as part of the channel, or
installed later either temporarily or permanently.
The depth of the flowing liquid is determined at a
specific point, or points, in the flume. The
measured depth, or depths, can then be used to
obtain the rate of flow of the liquid in the channel.
1.	Handbook of Hydraulics,
King, H. W., McGraw-Hill,
NY, 3rd Ed., 1939
2.	Water Measurement Manual,
US Dept. of the Interior,
Bureau of Reclamation,
Denver, CO, 2nd Ed., 1967
3.	Stevens Water Resources
Data Book, Leupold &
Stevens, Inc., Box 688,
Beaverton, OR 97005,
2nd. Ed., $4.00
Page No. 11-10
-------
EFFLUENT MONITORING PROCEDURE: Measurement of Flow in an Open Channel by Parshall Flume
EDUCATIONAL CONCEPTS - MATHEMATICS
Section II
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
A.l
Flows - Units of Expression
I. Flow, or Flow Rate, or Discharge.
A11 of these terms are commonly used to refer
to the quantity of liquid passing a point in
a certain time interval.
II. Quantity of liquid can be expressed in a number
of ways. Common units are the gallon (Gal) and
the cubic foot (cu.ft., ft.3). To change from
one of these measures to another, use the table
below:
Multiply by	To obtain
cu.ft. 7.5	Gal.
Gal.	0.134	cu.ft.
III. Flow is usually expressed in these units:
Gallons per minute (GPM)
Million gallons per day (MGD)
Cubic feet per second (cfs, Sec.-ft.)
To change from one of these units to another,
use this table:
Multiply
b*
To obtain
cfs
0.646
MGD
MGD
1.55
cfs
cfs
448.8
GPM
GPM
0.0022
cfs
MGD
694.4
GPM
GPM
0.00144
MGD
IV. Flow data is needed to calculate the quantity
of constituents discharged 1n a plant effluent.
Formulas are—
lb/day » MGD x mg/1 x 8.34
Kg/day * MGD x mg/1 x 3.78
Page No. 11-11
-------
£FFLUEflT MONITORING PROCEDURE: Measurement of Flow in an Open Channel by Parshall Flume
EDUCATIONAL CONCEPTS - MATHEMATICS
Section II
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
The head Ha is the vertical distance from the
crest of the flume (floor of the converging section)
to the liquid surface, at the stipulated point in the
converging section. Head is the corresponding
distance, as measured at the stipulated point in the
throat of the flume. Both of these measurements are
referenced to the same point, i.e., the elevation of
the crest of the flume. Consequently, all equipment
and devices used to measure these heads must also be
referenced to the crest elevation.
When a staff gage is used to obtain these heads,
it may be attached to the inside face of the flume,
or placed in a stilling well. In the former case,
only an approximate head determination is usually
possible, because of waves and rapid water level
fluctuations at the upstream gage, and turbulent
conditions at the downstream gage.
Determination of head using the staff gage is
illustrated below for the various conditions which
will be met.
Case I - Initial gage mark 0.00 ft.
The gage may be installed in either of three
positions, as shown in Fig. 1.
LIQUID
SURFACE
CREST,
0.30
0.20
1/W/V/V/V \rr7777
0.40
0.20
A.	B.	C.
FIG. 1 - STAFF GAGE SETTINGS
Page No. 11-12
-------
EFFLUENT MONITORING PROCEDURE: Measurement of Flow in an Open Channel by ParshaTI Flume
EDUCATIONAL CONCEPTS - MATHEMATICS
Section II
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
C.l
(cont.)
In "A", the bottom of the gage is set at crest
elevation. The intersection of the liquid surface
with the gage gives a direct reading of the head.
Here, the head is 0.30 ft.
In "B" the bottom of the gage is set some dis-
tance "d" above crest elevation. To obtain the
head, "d" must be added to the gage reading. For
example, if "d" in the Figure equals 0.25 ft., then
the head is 0.25 + 0.20, equals 0.45 ft.
In"C" the bottom of the gage is set some dis-
tance (say 0.20 ft) below crest elevation. This
must be subtracted from the gage reading to obtain
the head. Thus, 0.40-0.20 = 0.20 ft., which is the
head.
Case II - Initial gage mark other than 0.00 ft.
The mark at which the gage divisions start must
be taken into account in determining the head. For
example, if a gage section starting at 3.33 ft.
instead of 0.00 ft. is used, the calculations are
as follows for the three conditions shown in
Fig. 2 (which correspond to those of Fig. 1):
LIQUID
SURFACE
CREST
3.63
3.53
3.33

A//V/ 3.33	S7777777
A.
B.
C.
3.73
3.53
3.33
FIG. 2 - STAFF GAGE SETTINGS
In "A", head = 3.63-3.33 - 0.30 ft.
In "B", assuming that "d" ¦ 0.25 ft., head ¦
(3.53 + 0.25J-3.33 » 0.45 ft.
In "C", head - 3.73-3.53 = 0.20 ft.
I
Page No. 11-13
-------
TABLE 1
Page No. 11-14
FREE FLOW DISCHARGE TABLE FOR PARSHALL MEASURING FLUME
ft£C.-FT X	M.tt.D.
SEC-FT X 446.631 '6PM
HC AO

9"
It"
it"
M"

HEAD

9"
12"
18"
2«"
rt
IN.
sca-n
ft PM
M8.0
mere >ph
149.0
mcrr «.rm.I m.c 0
SCC-FT 8.P.M. ! M ao
SECfT
6PM
M. 6 0

FT IN
SCC FT; 9 P M
M.Q.O.
SECFT 6PM M 6 0
SCCFT 6PM
M 6 0
SCO-fT
G PM MCO
SECFT
G PM J M GO
aio
1*6
0.O5
2? 4
O032
O09| .10.4
.0560

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*'» 2
4650 • 6 960
Ofel
062
UT*«
094
4.2 0
6 60©
?.44
6460l 9300
iftft
©430! k.2«5
2.60 ; 1257
i.609
372
. IS70; 2 404
! nxo+i 463

22 4^
>23 14^
2.62
2.66
5 4: 74 26
3 437
9 5 1 j 5 26-1
4692
A940
7 045
097
999
•35 0
444 3
062-'
0.640
t.4©
¦-Si
664X1 9560
67ft.0j .9160
i-93
!9e
666.0' U241
&©9.0| 1260
2.67 [ >290
2 95 j 324
' 655
i.907
36!
1284
8a6
4 2 _ 894 _ 2 7C9
5 4* :.:6 2
8 25 ; 3^C3| 5 33?
; 7. :Q


3.91 T °55 : «i27
24 ,4
269 .297 86"
S 55 249 iS&"
836
e 4t
' 3"52_[ 5 4C3
5027! 7 24C
3.64

s.P2
4S~©
06SO
'.-55
S96.CT 1.0O2
7 03J 9" C, ,.3'2
302 ; '355
\ 952
4 0' i©00 2 592

^ D
293 3s 5 8*^4
4 3? 93^5 2
56? 252. 3 *"32
3SCO. :*468
' 3
5C "*2 7 304
Q65
-X.
04
466©
0612
'59
"714.0 1 02©
2 Oft! 934.0' 34S
309 ' 36->
».997
4 •. i 1 61S :656

26 5%
2 97 :333 92C
4 37 '96 ! 2V4
5 6°; <55.^ 3
' 6 258S • -; -3
53^ 26 : ^ 3 ~Z>Z
5 89 :644~3 8C"
5 56
"66^
! 364? j b5>3
' 5
5'62 ^7 433
"52 C 7",4?7'
Q66

•.07
460-0
0.692
1-63
732.0! 1.054
2 13 J 956 0;
317 : -42?
7 049
4.20' 665

1.27 5*
3c, ,150 ' ¦w
4 4:
= 2 Sfc.3
696
3892: S .^23
' 6
067
ft'-1*
!.-5
4937
'0.1: *
• 66i ",45.0j 1 073
2 l6i 976.C' !-409
3 24 '4 ^4
2 C94
4 301 .930
I 760
©44

.2ft
:9: o68
• ' ^ . 525 7562
0«,
8*fc
'.!2
5C30
0.724.
1.70 763.01 1 098
2.23 s 0 C
44
331 ! .466 » 2 >40
44Ci i915
4 53 2033 '
558 . 39S-6, 5 ~4C
' ' 9i 534 , "69
a69
6*
1.15
5<6 0
0.743
114! 7©» Oj 1 -25
2.26 < 024
1 414
3 39, 1522
2 191
4.so' 2020 j : 906
-.30' '.b\,
3 '2 j 400 | 2 0 -
4 59! 4 C6C "
596 26"»5 3 552
699 4T35 ' 5.6'0
'2 0j 53e6J 7 756
|aio
-ft
i.n
523.1
0756
.is! 1990I i.'5i
2.» '046
i 506
346 i553
2 236
460 i 20651 2 973

.31 i 15*
3.»6 1 '4'© I Z 04;
4 64 ; 2083 • 300O
6.C3 Z"-C i900
5-O-jU:oc! 5C5
2 2 ! S4^6 665
WATER * SEWAGE WORKS, REFERENCE AND DATA SECTION, 1954 P. R-277.
-------
EFFLUENT MONITORING PROCEDURE: 		 _r r1_ . «
	 Measurement of Flow in an Open Channel by Parshall Flume
EDUCATIONAL CONCEPTS - MATHEMATICS
Section II
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
With Ha determined, the flow can be obtained
from a Table such as that shown in Table 1. (This
shows the head-discharge relationships for flumes
ranging in size from 6 to 24 inches. Similar tables
for larger and smaller flumes will be found in
References 1,2, and 3).
The flow is obtained from this Table as follows
4. Water and Sewage Works,
Reference & Data Section,
1954, p. R-277
1.	Go vertically downward in the column titled
"Head" until you reach the value for the Ha
measured. Note that values of Head in this
column are given both in feet, and in inches
corresponding to the foot values.
2.	Proceed horizontally to the right until you
reach the columns for the throat width of
flume in use.
3. Read the flow in the units to be reported.
The flow is given in three units in this
Table: sec.-ft. (cubic feet per second);
GPM (gallons per minute), and M6D (million
gallons per day). Example:
For a 12" flume, with Ha = 0.86 ft.
1.	Locate 0.86 ft. in the "Head" column.
2.	Go over horizontally to the right to the
columns under the "12" throat width.
3.	Read flow: 3.18 sec.-ft., or
1427 GPM, or
2.06 MGD
Page No. 11-15
-------
EFITUI.NT MONITORING PROCCfJlJR
Measurement of Flow in an Open Channel by Parshall Flume
EDUCATIONAL CONCEPTS - MATHEMATICS
Section II
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
D.l
A float gage is shown in Fig. 3,
stilling well for measurement of H
installed in a
To illustrate
the calculation of Ha it is assumed that the floor
of the stilling well is at the same elevation as the
INDEX
PULLEY
FLOAT
1 FOOT
FIG.3 - FLOAT GAGE INSTALLATION
crest of the flume, and that the liquid is flowing
in the flume with a depth of one foot. The float
will, of course, be resting on the surface of the
liquid in the well, and it is also assumed that for
the condition illustrated the tape division opposite
the float index reads 8-1/2 feet.
With the specific relations established for
this one condition, the gage can now be "zeroed" so
that Ha can be obtained for any other condition, as
follows:
(a) A reading of 8-1/2 feet on the tape corre-
sponds to an Ha of one foot. Consequently,
a reading of 7-1/2 feet on the tape corre-
sponds to an Ha of zero feet, or to the
crest elevation.
(bJ Therefore Ha can be obtained for any depth
of flow by subtracting 7-1/2 feet from the
observed tape division opposite the index.
The following points should be noted in connec-
tion with this procedure:
(a) If the elevation of the index is changed,
the gage must be re-zeroed.
Page No. 11-16
-------
EFFLUENT MONITORING PROCEDURE;: Measurement of Flow in an Open Channel by Parshall Flume
EDUCATIONAL CONCEPTS - MATHEMATICS
Section II
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
(b)	If the position of the tape on the pulley is
changed, the gage must be re-zeroed.
(c)	The tape must be installed so that the
numerical value of the tape reading
increases as the depth of flow increases.
Hfc can be obtained with the float gage in the
same manner as described above for Ha.
Discharge through the flume is not reduced from
the free-flow value until the percent submergence
equals or exceeds the following values:
60% for 6-inch and 9-inch flumes
70% for 1 foot to 8-foot flumes
When the submergence reaches these values, a
corrected flow must be calculated in the following
manner:
For 6-inch and 9-inch Flumes
The corrected flow can be obtained directly
from Fig. 4 for a 6-inch flume, and from Fig. 5 for
a 9-inch flume. Example: For a 6-inch flume,
Ha = 1.0 ft., Hb = 0.8 ft.
% Submergence = x 100 = 80%
Refer to Fig. 4. On "Percent of submergence"
scale on left-hand side, go up to the "80"
value. Move to the right along the "80" line to
where it intersects the "Ha = 1.0 feet" curve.
Drop vertically from the point of intersection
to the "Discharge, Second-feet" scale along the
bottom of the chart.
Read 1.7 - this is the discharge in cubic feet
per second through the flume. Convert flow to
other units if desired.
Exactly the same procedure would be followed for a
9-inch flume, using Fig. 5.
Ref. 1
Page No. 11-17
-------
EFFLUENT MONITORING PROCEDURE: Measurement of Flow in an Open Channel by Parshall Flume
UPSTREAM HfAD Ho, MET
Ui
Z
o
IX
o
o
a
a
4 O
DISCMARGE, second-feet
FIG. 4 - Diagram for determining rate of submersed flow for a 6-inch Parshall flume. 103-D-897. (Courtesy U.S.
Soil Conservation Service.)
'PSTREAVI Hi AP ho, f f fr T
60
6?
68
•70
i-
uj 74
O
5'76
2 re
96
*0
6 0
55
DISCHARGE, SECOND-FEET
4 0
I 5
DISCHARGE
FIG. 5—Diagram for determining rate of submerged How lor a 9-inch Parshall flume. 103-D-898. (Courtesy U.S.
Soil Conservation Service.)
Page No. 11-18
-------
EFFLUENT MONITORING PROCEDURE; Measurement of Flow in an Open Channel by Parshall Flume
EDUCATIONAL CONCEPTS - MATHEMATICS 	Section II
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
E.3.2
(cont.)
For Flumes 1 foot to 8 feet wide
Use Fig. 6. This provides a correction factor
to be applied to the discharge obtained using Ha
and Table 1, the free-flow discharge table. For
flumes larger than 1 foot a second correction, using
a "multiplying factor" is necessary. Example 1.
For a 1-foot flume,
Hfa = 0.8 ft., Ha = 1.0 ft.
Submergence = x 100 = 80%
1.0
Refer to Fig. 6. On "Upstream Head Hfl" scale
at left-hand side, go up to 1.0 ft. Move to
the right along the "1.0 ft." line to where
it intersects the "80% Submergence" curve.
Drop vertically from the point of intersection
to the "Correction, second-feet" scale at the
bottom of the chart. Read "0.35 sec.-ft."
Refer to Table 1. For a 12-inch flume with
Ha = 1.0 ft., discharge is 4.00 sec.-ft. But
the actual discharge will be less than this,
since submergence exceeds 70%. To get actual
discharge, subtract correction obtained from
Fig. 6. Then the discharge is 4.00-0.35
= 3.65 sec.-ft.
Note that "Multiplying Factor" is 1.0, so the
correction factor obtained from Fig. 6 1s
used directly.
Example 2
For a 24-inch flume,
Hb = 1.23 ft., Ha = 1.30 ft.
Submergence = 1^1 x 100 = 95%
1.30
Refer to Fig. 6. On left-hand scale go up to
1.30 ft., which 1s the Upstream Head Ha.

Page No. 11-19
-------
Page No. 11 -20

SIZE OF FUME
W, FEET
MULTIPLYING
FACTOR. M
¥A
NOTE Corr#ction is ui«d dir»ctly for I
foot fluw»s For lorjtr mti *h§
corr«etion «quol* th« volu* from th»
chart multiplnd by (>:• foctor, M
22* *
06 ,07 .06.09.10 .12 .14 J6 .18.20 .25 .30 ,35 .40 .45.50 . 60 .70 .80.90 1,0 1.2 L4 1,6 13 2,0 2.5
CORRECTION, SECOND-FEET
FIG. 6—Diagram for determining correction to be subtracted from free-discharge flow to obtain rate of submerged flow
through Parshall flumes 1 to 8 feet wide. 103-D-875. (Courtesy U.S. Soil Conservation Service.)
-------
EFFLUENT MONITORING PROCEDURE: Measurement of Flow in an Open Channel by Parshall Flume
EDUCATIONAL CONCEPTS - MATHEMATICS
TRAINING GUIDE MOTE
REFERENCES/RESOURCES
E.3.2
(cont.)
Proceed horizontally to the right along the
"1.30 ft." line. The-point of intersection of
this line with the "95% Submergence" curve is to
be located. Since no curve is drawn on Fig. 6
for this value of submergence, mentally locate a
point on the "1.30 ft." line which is midway
between the "94% submergence" and the "96% sub-
mergence" curves.
Drop vertically from this point to intersect the
"Correction, second-feet" scale at the bottom of
the chart. Read "2.7".
From the Table at right side of chart on Fig. 6,
read the "Multiplying Factor" for a 24-inch
flume. This factor is 1.8.
Multiply. 2.7 x 1.8 = 4.9 sec.-ft. This is the.
correction factor to be used in this case.
From Table 1 obtain free-tlow discharge of 12.0
sec.-ft. for a 2-foot flume with Ha = 1.30 ft.
Subtract correction factor from this free-flow
value to obtain discharge with this degree of
submergence.
Discharge = 12.0 - 4.9 = 7.1 sec.-ft.

Page No. 11-21
-------
n_ri-llENT j'lOjj IJj' KING PROCEDURE: Measurement of Flow in an Open Channel by Parshall Flume
EDUCATIONAL CONCEPTS - SCIENCE
Section III
TRAINING GUIDE MOTE
REFERENCES/RESOURCES
The Parshall Flume is intended for use as an
in-line structure in an open channel where reason-
ably smooth flow, uniformly distributed across the
cross-section-, is the normal condition.
A good degree of accuracy cannot be maintained
if poor approach conditions exist in the approach
channel. Experience has shown that Parshall Flumes
should not be placed at right angles to flowing
streams unless the flow is effectively straightened
and uniformly redistributed before it enters the
flume. Surges and waves of any appreciable size
should be eliminated.
The liquid should enter the converging section
reasonably well distributed across the entrance
width, and the flowlines should be essentially
parallel to the flume centerline. Flow at the flume
entrance should be free of "white" water and free
from turbulence in the form of visible surface
boils. Only then can the flume measure flow as
intended.
The velocity of flow through the flume will
generally be sufficiently great to virtually
eliminate any deposition of sediment within the
structure. If any such build-up is observed,
however, it should be eliminated. Deposits should
also be removed from the channel upstream and down-
stream of the flume.
The flow condition can be determined from
measurements of Ha and Hu. Generally, however,
these heads do not have to be measured—the condition
of flow through the flume can usually be determined
by visual observation.
Three flow conditions through the flume are
shown in Fig. 7.
Hf
Hfc
J
Flow
f
- -Q .
FIG. 7 - FLOW CONDITIONS
Page No. 11-22
-------
EFFLUENT MONITORING PROCEDURE: 		 ri	„	. . _
	 Measurement of Flow in an Open Channel by ParshaU Flume
EDUCATIONAL CONCEPTS - SCIENCE	Section III
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
B.1.4
(cont.)
In flow condition 2, there is a drop in the
elevation of the liquid surface, followed by an
abrupt rise in the throat*. This phenomenon is
referred to as a hydraulic jump or standing wave.
When the hydraulic jump is present, free-flow
conditions exist.
In flow condition 1, there is a substantial and
smooth drop in the elevation of the liquid surface
as it passes through the throat and the diverging
section of the flume. Free-flow conditions exist.
A hydraulic jump will be observed downstream of
the flume.
Flow condition 3 illustrates the configuration
of the liquid surface for submerged-flow conditions.
Sometimes a series of waves or ripples will be noted
in the transition area between the upstream and
downstream liquid elevations. These also indicate
that submerged-flow conditions exist.
Flumes used in treatment plants are selected
to operate under free-flow conditions over the
range of flows handled at the plant. The existence
of a submerged-flow condition would therefore be
most unusual, and might be due either to the flume
being too small, or to some obstruction in the
channel downstream of the flume which is raising the
water level. In any case, it is important to
determine the reason for a submerged flow Condition,
and take the appropriate steps to return the flume
to free-flow operation.

B.1.5
For a stilling well to function properly, the
opening or pipe between the well and the flume must
be kept free of deposits or materials which would
interfere with the free movement of liquid. This
should be checked occasionally, and any such Inter-
ferences removed by flushing with clean water, or
by some other suitable procedure. Deposits or
floating materials in the well should also be
removed to maintain accurate head measurement.

Paqe No. 11-23
-------
EFFLUENT MONITORING PROCEDURE: Measurement of Flow in an Open Channel by Parshall Flume
FIELD & LABORATORY EQUIPMENT	Section V

TRAINING GUIDE NOTE
REFERENCES/RESOURCES
P.3
Two drawings of a Parshall Flume are shown in
Fig. 8. The top drawing shows the appearance of the
flume when viewed from above. The drawing labeled
"Section L-L" is the way the flume looks when viewed
from the side, along the line marked "L-L" in the
top drawing.

Converging Section	Diverging
Section
Throat
Section
Flow
PLAN
Wingwall
Stilling Wells
Water Surface
Flow
Crest
SECTION L-l
Fig. 8. PARSHALL FLUME
The flume structure proper consists of three
sections:
1.	A converging section
2.	A throat section
3.	A diverging section
(continued)
Page No. 11-24
-------
EFFLUENT MONITORING PROCEDURE: Measurement of Flow in an Open Channel by Parshall Flume
FIELD & LABORATORY EQUIPMENT
Section v
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
A.3 (cont'd.)
A.4
Wing walls are shown in Fig. 8 immediately up-
stream and downstream of the flume. In situations
where the channel in which the flume is located is
wider than the beginning of the converging section
and the end of the diverging section of the flume,
these wing walls provide a gradual transition in
width of the flowing liquid. Their function is to
ensure proper approach and "getaway" conditions to
avoid measurement difficulties. Curved winqwalls.are
preferred over straight 45 walls, althouqh anv
arrangement that achieves uniformity and smoothness
in the flow is acceptable.
Flumes can be made of concrete, galvanized steel,
plastic, wood, or other suitable material. Flume
must be built to specific dimensions, for which
tables are available (page 47, Ref. 2), and close
tolerances for accurate performance. The floor of
the converging section (flume crest) must be level
if the flume is to operate properly.
Throat Width
Distance between the walls of the throat section.
Flume Size
Flumes are designated as to size by the throat width,
as a "6-1nch flume," a "10-foot flume," etc.
Flume Crest
Floor of the converging section. Sometimes indicated
as the junction point of the floor of the converging
section with the throat section.
Crest Elevation
Elevation of the floor of the converging section.
Upstream Head (Ha)
Depth of liquid in the converging section, measured
at a point two-thirds of the section length upstream
from the throat (see Fig. 8).
Downstream Head (Hb)
Depth of liquid over the flume crest, measured at a
specific point in the throat section. For flumes
considered in this guide the point of measurement
is 2 inches upstream of the beginning of the diverging
section. Because of turbulence In the throat section,
1t 1s often impossible to determine the head accu-
rately with a staff gage, and a stilling well should
be provided. The connection between the throat and
I	(continued)
Page No. 11-25
-------
EFFLUENT MONITORING PROCEDURE: Measurement of Flow in an Open Channel by Parshall Flume
FIELD & LABORATORY EQUIPMENT
Section V
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
A.4 (cont'd.)
A.5
stilling well must be located two inches upstream
from the low point in the floor (dimension X. Fig. 8)
and 3 inches above it (dimension Y, Fig. 8).
Stilling Well (Float Well)
A chamber connected through a small opening to the
liquid flowing in an open channel. Waves and surges
occurring in the flowing liquid will not appear in
the well. Liquid level in the well will follow all
the steady fluctuations of the flowing liquid. The
well must have a bottom and be practically water
tight except for the liquid inlet.
Free Flow
A flow condition in which liquid passing over the
crest of the flume is not impeded or slowed by
downstream conditions.
Submerged Flow
In most installations, when the discharge through
the flume is increased above a certain critical value
the resistence to flow in the downstream channel
becomes large enough to reduce the velocity, increase
the flow depth and cause a back water effect at the
flume, in which the flow is retarded and discharge
reduced.
Submergence
The ratio H^, usually expressed as a percentage.
H_
The Parshall Flume is a specially-shaped flow
section, so constructed and installed that the rate
of flow through it depends only on its size (throat
width), and the depth of liquid over the crest.
Discharge through the flume can occur for two con-
ditions of flow:
1.	Free Flow, in which the discharge depends
only on the upstream head Ha. When free-
d
flow conditions exist, discharge through the
flume can be obtained by measuring the up-
stream head only.
2.	Submerged Flow, in which the discharge 1s
reduced due to the effect of the depth of
liquid downstream of the flume. In this
case it 1s necessary to measure both the
Page No. 11-26
upstream head and the downstream head
in order to obtain the discharge.
-------
EFFLUENT MONITORING PROCEDURE: Measurement of Flow in an Open Channel by Parshall Flume
FIELD & LABORATORY EQUIPMENT
Section V
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
A.6
A.7
A staff gage (Fig. 9) is a graduated scale,
usually installed vertically, for obtaining liquid
depth, or head. An observer notes the scale divi-
sion at which the liquid surface intersects the
9~i
«J
e
s-i
4-1
3-1
8-=
FIG. 9 - STAFF
GAGE SECTION
The head and discharge can then
gage (gage height),
be calculated.
Commercially-available gages are made of 18-gage
metal coated with a substantial thickness of porce-
lain enamel. The face of the gage is white; numerals
and graduations are black. Gages are available in
several styles; in widths from 2-1/2 to 4 inches, in
lengths from 1 to 5 feet. A gage divided in metric
units is also commercially available.
A float gage (Fig. 10) is a means of continu-
ously indicating liquid levels. It consists of a
metal float, a pulley mounted on a standard, and a
counterweight. A graduated stainless steel tape 1s
(Fig. 10)
attached to the float and connected at the other end
to the counterweight. The float follows the rise and
(continued)
Page No. 11-27
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EFFLUENT MONITORING PROCEDURE: Measurement of Flow in an Open Channel by Parshall Flume
FIELD & LABORATORY EQUIPMENT	Section V

TRAINING GUIDE NOTE
REFERENCES/RESOURCES
A.7 (cont'd.)
fall of the liquid surface and the level can be read
from the tape and a pointer or reference nark. Tapes
are available in selected lengths, and are graduated
either in feet, tenths and hundredths for English
measurements, or meters, decimeters and centimeters
for metric measurements.
The float gage is used extensively as a refer-
ence gage in stilling wells to check the accuracy
of automatic head or flow recording devices.

Page No. 11-28
-------
A PROTOTYPE FOR DEVELOPMENT OF
ROUTINE OPERATIONAL PROCEDURES
for the
MEASUREMENT OF FLOW IN AN OPEN CHANNEL BY
SHARP-CRESTED WEIR
as applied in
WASTEWATER TREATMENT FACILITIES
and in
MONITORING OF EFFLUENT WASTEWATERS
Developed by the
National Training and Operational Technology Center
Municipal Operations and Training Division
Office of Water Program Operations
U.S. Environmental Protection Agency
EN.FM.EMP.la.8.76
Page No. 12-1
-------
EFFLUENT MONITORING PROCEDURE:
Measurement of Flow in an Open Channel by
Sharp-Crested Weir
This Procedure was developed by:
NAME	Charles E. Sponagle
ADDRESS	EPA, OWPO, NTOTC, Cincinnati, Ohio 45268
POSITION Sanitary Engineer-Instructor
EDUCATION AND TECHNICAL BACKGROUND
B.C.E. - Manhattan College, 1943
M.S. in C.E. - University of Minnesota, 1948
Professional registration, New York State
With Federal Water Pollution Control Program since
1948, with various assignments at Program Headquarters,
Regional Offices, and Field Stations, including positions
as
Staff Engineer, then Chief, Water Quality Section,
Denver Regional Office
Staff Engineer, then Regional Construction Grants
Program Director, Denver Regional Office
Regional Construction Grants Program Director,
Cincinnati Regional Office
Director, Colorado River Basin Water Quality
Control Project, Denver, Colorado
Industrial Wastes Consultant, Technical Advisory
and Investigations Branch, Cincinnati, Ohio
Participation in and Direction of numerous in-plant
industrial waste surveys and stream studies in New
York, Colorado, New Mexico, Maine, Utah
With National Training Center September 1969 to date.
Paqe No. 12-3
-------
EFFLUENT MONITORING PROCEDURE: Measurement of Flow in an Open Channel by
Sharp-Crested Weir
1. Objective:
The student will be able to make an acceptable measurement of flow rate
in an open channel by means of a preinstalled sharp-crested weir and
vertical staff gage or a float gage.
2. Brief Description of Procedure:
The depth of liquid producing flow over a weir is measured. This measure-
ment is used to obtain the rate of flow in the channel at the time the
observation was made.
General Description of Equipment used in the Procedure:
1.	A Weir over which the liquid flows.
2.	Means for visually observing depth of liquid above the weir crest,
such as a staff gage or a float gage.
Page No. 12-4
-------
EFFLUENT MONITORING PROCEDURE: Measurement of Flow in an Open Channel
by Sharp-Crested Weir
Page No. 12-6
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
C. Flow Measurement
Using Staff Gage
1.	Detenriination of
Head
2.	Determination of
Flow Rate
1.	Read gage division at which
liquid surface intersects
gage.
2.	Calculate head on weir.
1. Use appropriate weir table.
la. To nearest division.
lb. This reading should be made at a distance at
least 2.5 H upstream of the weir.
2a. From staff gage reading.
2b. Should not be less than 0.2 feet.
II.C.l (p. 10)
II.C.2 (p. 10)
D. Flow Measurement
Using Float Gage
1.	Determination of
Head
2.	Determination of
Flow Rate
1.	Read tape division opposite
index on float gage.
2.	Calculate head on weir.
1. Use appropriate weir table.
la. To nearest division
lb. This reading should be made at a distance at
least 2.5 H upstream of the weir.
2a. From float gage reading
2b. Should not be less than 0.2 feet.
II.D.l (P- 15)
II.C.2 (p. 15)
-------
EFFLUENT MONITORING PROCEDURE: Measurement of Flow in an Open Channel
" ""	'	by Sharp-Crested Weir
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
A. Basic Elements
1.	Units of Flow
Measurement
2.	Description of
Process
3.	Definitions
4.	Types of Weirs
5.	Staff Gage
6.	Float Sage
II.A.1 (p.	9)
I.A.2 (p.	8)
V.A.3 (P ¦	16)
V.A.4 (P-	16)
V.A.5 (p.	18)
V.A.6 (p.	18)
B. Preparation for
Measurement
1. Physical
Conditions
1. Inspect weir bulkhead.
2. Inspect weir plate.
3. Inspect nappe.
4. Inspect approach channel
5. Allow flow to stabilize.
la. No leakage.
lb. Bulkhead vertical.
lc. Bulkhead perpendicular to direction of flow.
2a. Crest horizontal
2b. Crest at zero gage elevation
2c. No nicks or dents
2d. No clinging debris or build-up of grease, etc.
3a. No submergence.
3b. Springs clear of downstream side of weir plate.
4a. No large submerged or floating objects.
4b. No excessive sediment deposits.
5a. Undisturbed flow condition.
V.B.I .1 (p.	19)
V.B.I.2 (P-	19)
V.B.I.3 (p.	20)
V.B.I .4 (P-	20)
V.B.I.5 (p-	21 )
Page No. 12-5
-------
EFFLUENT MONITORING PROCEDURE:
Measurement of Flow In an Open Channel
by Sharp-Crested Weir
TRAINING GUIDE
SECTION	TOPIC
I*	Introduction
II*	Educational Concepts - Mathematics
III	Educational Concepts - Science
IV	Educational Concepts - Communication
V*	Field & Laboratory Equipment
VI	Field & Laboratory Reagents
VII	Field & Laboratory Analysis
VIII	Safety
IX	Records & Reports
~Training guide materials are presented here under the headings marked *.
These standardized headings are used throughout this series of procedures.
Page No. 12-7
-------
EFFLUENT MONITOR I * IH PROCEDURE: Measurement of Flow in an Open Channel
by Sharp-Crested Weir
Introduction			Section I
TRAINIKG GUIDE NOTE
REFERENCES/RESOURCES
A.2
Flow of a liquid in an open channel can often be
conveniently and accurately measured by means of a
sharp-crested weir installed in the channel. For a
weir of specific size and shape with free-flow
steady-state conditions and proper approach con-
ditions, only one depth of liquid can exist upstream
of the weir for a given flow. The flow is determinec
by measuring the vertical distance from the crest of
the weir plate to the upstream liquid surface and
then using a weir formula or weir table. The weir
must have a standard shape and dimensions, and be
installed so that the system performs in a standard
manner.
1.	Handbook of Hydraulics
King, H.W., McGraw-Hill,
NY, 3rd Ed. 1939
2.	Water Measurement
Manual, US Dept. Interior,
Bur. Reclamation, Denver,
CO, 2nd Ed. 1967
3.	Stevens Water
Resources Data Book.
Leupold & Stevens,
Inc., Box 688,
Beavertoh, Oregon,
97005. 2nd Ed.
$4.00
Page No. 12-8
-------
EFFLUENT MONITORING PROCEDURE: Measurement of Flow 1n an Open Channel
by Sharp-Crested Weir
Educational Concepts - Mathematics
Section II
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
A.l
Flows - Units of Measurement
I. Flow, or Flow Rate, or Discharge.
All of these terms are commonly used to refer
to the quantity of liquid passing a point in a
certain time interval.
Quantity of liquid can be expressed in a
number of ways. Common units are the gallon
(Gal) and the cubic foot (cu. ft., ft.3). To
change from one of these measures to another,
use the table below:
II.
Multiply by To obtain
cu. ft.
Gal.
7.5
0.134
Gal.
cu. ft.
III. Flow is usually expressed in these units:
Gallons per minute (GPM)
Million gallons per day (MGD)
Cubic feet per second (cfs)
To change from one of these units to another,
use this table:
Multiply
hx.
To obtain
cfs
0.646
MGD
MGD
1.55
cfs
cfs
448.8
GPM
GPM
0.0022
cfs
MGD
694.4
GPM
GPM
0.00144
MGD
IV. Flow data is needed to calculate the quantity
of constituents discharged in a plant effluent.
Formulas are—
lb/day = MGD x mg/1 x 8.34
Kg/day = MGD x mg/1 x 3.78
Page No. 12-9
-------
EFFLUENT MONITORING PROCEDURE: Measurement of Flow in an Open Channel
by Sharp-Crested Weir
Educational Concepts - Mathematics
Section II
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
C.I
The head on the weir is calculated from the staff
gage reading. Either of two conditions may exist,
depending on zero gage elevation:
Case I - Zero gage elevation is at "0" on the gage
LIQUID
SURFACE 3
WEIR
CREST
FIG. 1 - HEAD ON WEIR
The head on the weir corresponds to the gage division
intersected by the surface of the liquid. In the
above diagram H=2 feet.
Case II - Zero gage elevation is at some gage •
division other than "0"
The diagram below illustrates this case when the
1-foot division on the gage is at the same elevation
as the weir crest.
LIQUID
SURFACE
GAGE
WEIR
CREST
FIG. 2 - HEAD ON WEIR
Since the head on the weir is the difference between
zero gage elevation and the gage division intersected
by the liquid surface, H=3-l=2 feet.
Having determined the head on the weir, the flow
rate can be obtained from a weir table. The proper
table for the type of weir in use must be selected.
The use of weir tables is shown below.
Page No. 12-10
-------
EFFLUENT MONITORING PROCEDURE: Measurement of Flow in an Open Channel
by Sharp-Crested Weir
Educational Concepts - Mathematics
Section II
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
C.2 (Cont'd)
Use of weir tables.
1.	90° V-notch weir - Table I
This table lists flows corresponding to weir heads
ranging from 0.10' to 2.09'. The flow for any head
in this range can be read directly from the table.
Example: For H=0.65'
At intersection of values of 0.60 in left-hand
head column and 0.05 in top column, read
Q=0.852 cfs or 0.550 MGD
2.	Standard Contracted Rectangular Weir - Table II
Flows are given for various heads and for weirs
having different crest lengths.
Example: Weir crest=3'
H=0.26'
Read from table 0=1.30 cfs or 0.84 MGD
3.	Standard Suppressed Rectangular Weir •- Table III
The format of this table differs from that of
Table II, 1n that the flow is given per foot of
weir crest length. Values obtained from the
table must therefore be multiplied by the
crest length of the weir to obtain the total
flow.
Example: Weir crest length=10'
H=1.0'
From table, Q=3.33 cfs or 2.15 MGD
This is the flow per foot of weir length; there-
fore the total flow over the weir 1s
Q=3.33 x 10=33.3 cfs or 21.5 MGD
Ref. 3
Ref. 3
Ref. 3
Page No. 12-11
-------
EFFLUENT MONITORING PROCEDURE: Measurement of Flow in an Open Channel
by Sharp-Crested Weir
Page No. 12-12
TABLE 1
DISCHARGE OF 90° V-NOTCH WEIRS
FORMULA CFS= 2.50 H 5/2 MGD = CFS x 646317
Head
Ft.
.00
.01
.02
.03
.04
.05
.06
.07
.08
.09
CFS
MGD
CFS
MGD
CFS
MGD
CFS
MGD
CFS
MGD
CFS
MGD
CFS
MGD
CFS
MGD
CFS
MGD
CFS
MGD
0.1
.008
.005
.010
.006
.012
.008
.015
.010
.018
.012
.022
.014
.026
.017
.030
.019
.034
.022
.039
.025
0.2
.045
.029
.051
.033
.057
.037
.063
.041
.071
.046
.078
.050
.086
.056
.095
.061
.104
.067
.113
.073
0.3
.123
.079
.134
.087
.145
.094
.156
.101
.169
.109
.181
.117
.194
.1 25
.208
.134
.223
.144
.237
.153
0.4
.253
.164
.269
.174
.286
.185
.303
.196
.321
.207
.340
.22(1
.359
.232
.379
.245
.399
.258
.420
.271
0.5
.442
.286
.464
.300
.487
.315
.511
,330
.536
.346
.561
363
.587
3 79
.613
.3%
.640
.414
.668
.432
0.6
.697
.450
.727
.470
.757
.489
.788
.509
.819
.529
.852
.55!
.885
5 7 2
.919
.594
.953
.616
.989
.639
0.7
1.03
.662
1.06
.686
1.10
.711
1.14
.736
1.18
.761
1.22
.787
1.26
814
1.30
.841
1.34
.868
1.39
.896
0.8
1.43
.925
1.48
.954
1.52
.984
1.57
1.01
1.62
1.05
1.67
1.08
1.71
Ml
1.76
1.14
1.82
1.18
1.87
1.21
0.9
1.92
1.24
1.97
1.27
2.03
1.31
2.09
1.35
2.14
1.38
2.20
1.42
2.26
1.46
2.32
1.50
2.38
1.54
2.44
1.58
1.0
2.50
1.62
2.56
1.65
2.63
1.70
2.69
1.74
2.76
1.78
2.82
1.82
2.89
1.87
2.96
1.91
3.03
1.96
3.10
2.00
1.1
3.17
2.05
3.25
2.10
3.32
2.15
3.39
2.19
3.47
2.24
3.55
2.29
3.62
2.34
3.70
2.39
3.78
2.44
3.86
2.49
1.2
3.94
2.55
4.03
2.60
4.11
2.66
4.19
2.71
4.28
2.77
4.37
2.82
4.46
2.88
4.54
2.93
4.63
2.99
4.73
.3.06
1.3
4.8;
3.12
4.91
3.17
5.00
3.23
5.10
3.30
5.20
3.36
5.29
3.42
5.39
3.48
5.49
3.5 5
5.59
3.61
5.69
3.68
1.4
5.80
3.75
5.90
3.81
6.01
3.88
6.11
3.95
6.22
4.02
6.33
4.09
6.44
4.16
6.55
4.23
6.66
4.30
6.77
4.38
1.5
6.89
4.45
7.00
4.52
7.12
4.60
7.24
4.68
7.36
4.76
7.48
4.83
7.6d
¦4.91
7.72
4.99
7.84
5.07
7 97
5.15
1.6
8.10
5.24
8.22
5.31
8.35
5.40
8.48
5.48
8.61
5.56
8.74
5.65
8.88
5.74
9.01
5.82
9.15
5.91
9.28
6.00
1.7
9.42
6.09
9.56
6.18
9.70
6.27
9.84
6.36
9.98
6.45
10.1
6 55
10.3
6.64
10.4
6.73
10.6
6.83
10.7
6 93
1.8
10.9
7.03
11.0
7.12
11.2
7.22
11.3
7.32
11.5
7.42
11.6
7.52
1 1.8
7.63
11.9
7.72
12.1
7.83
12.3
7.94
1.9
12.4
8.04
12.6
8.14
12.8
8.25
12.9
8.36
13.1
8.47
13.3
8.58
13.5
8.69
13 6
8.80
13.8
8.91
14.0
9,o3
2.0
14.1
9.14
14.3
9.26
14.5
9.37
14.7
9.49
14.9
9.60
15.0
9.72
15.2
9.84
15.4
9.96
15.6
10.1
15.8
10.2
IEUPOID 4 STEVENS, INC
STEVENS WATER RESOURCES DATA, BOOK 2nd ED
-------
EFFLUENT MONITORING PROCEDURE: Measurement of Flow in an Open Channel
by Sharp-Crested Weir
TABLE 2
FLOW THROUGH RECTANGULAR WEIRS
WITH END CONTRACTIONS
Formula CFS = 3.33(L-0.2H)H3/2	MGD = CFS X .646317
Head
Ft.
LKNGTH 01 WFIR CRliST IN I L L
T
1
1 Vi
2
3
4
5
(TS
MGD
CI s
MGD
OS
MGD
CI'S
MGD
CIS
MGD
CI S
MGD
.01
.003
.002
.005
.003
.007
.005
.010
.006
.013
.008
.017
.011
.02
.009
.006
.014
.009
.019
.012
.028
.018
.038
.025
.047
.030
.03
.017
.011
.026
.017
.035
.023
.052
.034
.069
.045
.086
.056
.04
.026
.017
.040
.026
.053
.034
.080
.052
.106
.069
.133
.086
.05
.037
.024
.055
.036
.074
.048
111
.072
.149
.096
.186
.120
.06
.048
.031
.073
.047
.097
.063
.146
.094
.195
.126
.244
.158
.07
.061
.039
.092
.059
.122
.079
.184
.119
.246
.159
.307
.198
.08
.074
.048
.112
.072
.149
.096
.225
.145
.300
.194
.376
.243
.09
.088
.057
.133
.086
.178
.1 15
.268
.173
.358
.231
.448
.290
.10
.103
.067
.156
.101
.209
.135
.314
.203
.419
.271
.524
.339
.11
.119
.077
.180
.1 16
.240
.155
.362
.234
.483
.312
.605
.391
.12
.135
.087
.204
.132
.274
.177
.412
.266
.550
.355
.689
.445
.13
.152
.098
.230
.149
.308
.199
.464
.300
.620
.401
.776
.502
.14
.170
.1 10
.257
.166
.344
.222
.518
.335
.693
.448
.867
.560
.15
.188
.122
.284
.184
.381
.246
.575
.372
.768
.496
.961
.621
.16
.206
.133
.313
.202
.419
.271
.633
.409
.846
.547
1.059
.684
.17
.225
.145
.342
.221
.459
.297
.692
.447
.926
.598
1.159
.749
.18
.245
.1S8
.372
.240
.499
.323
.754
.487
1.008
.651
1.262
.816
.19
.265
.171
.404
.260
.541
.350
.817
.528
1.093
.706
1.368
.884
.20
.286
.185
.435
.281
.584
.377
.882
.570
1.179
.762
1.477
.955
.21
.307
! 198
.468
.302
.627
.405
.948
.613
1.268
.820
1.589
1.027
.22
.329
.213
.501
.323
.672
.434
1.016
.657
1.359
.878
1.703
1.101
.23
.350
.226
.534
.345
.718
.464
1.085
.701
1.452
.938
1.820
1.176
.24
.373
.241
.568
.367
.764
.494
1.156
.747
1.547
1.000
1.939
1.253
• .25
.395
.25S
.604
.390
.812
.525
1.228
.794
1.644
1.063
2.060
1.331
.26
.419
.271
.639
.413
.860
.556
1.301
.841
1.743
1.127
2.184
1.412
.27
.442
.286
.676
.437
.909
.588
1.376
.889
1.844
1.192
2.311
1.494
.28
.466
.301
.712
.460
.959
.620
1.453
.939
1.946
1.258
2.439
1.576
.29
.490
.317
.750
.485
1.010
.653
1.530
.989
2.050
1.325
2.570
1.661
.30
.514
.332
.788
.509
1.062
.686
1.609
1.040
2.156
1.393
2.703
1.747
.31
.539
.348
.827
.535
I.I 14
.720
1.689
1.092
2.263
1.463
2.838
1.834
.32
.564
.365
.866
.560
1.167
.754
1.770
1.144
2.373
1.534
2.975
1.923
.33
.590
.381
.905
.585
1.221
.789
1.852
1.197
2.483
1.605
3.115
2.013
.34
.615
.397
.945
.611
1.275
.824
1.936
1.251
2.596
1.678
3.256
2.104
.35
.641
.414
.986
.637
1.331
.860
2.020
1.306
2.710
1.752
3.399
2.197
IEUPOLD & STEVENS, INC.
STEVENS WATER RESOURCES DATA BOOK. 2nd ED.
-------
Page No. 12-14
EFFLUENT MONITORING PROCEDURE: Measurement of Flow in an Open Channel
by Sharp-Crested Weir
TABLE 3
FLOW PER FOOT OF LENGTH THROUGH RECTANGULAR
WEIRS WITHOUT END CONTRACTIONS
Formula CFS = 3.33 L H3/2	MGD = CFS x .646317
Head
Ft.
.00
.01
.02
• .03
.04
.05
.06
.07
.08
.09
CFS
MGD
CFS
MGD
CFS
MGD
CFS
MGD
CFS
MGD
CFS
MGD
CFS
MGD
CFS
MGD
CFS
MGD
CFS
MGD
0.0
.00
.00
.00
.00
.01
.01
.02
.01
.03
.02
.04
.03
.05
.03
.06
.04
.08
05
.09
.0(5
.1
.11
.07
.12
.08
.14
.09
.16
.10
.17
.11
.19
.12
.21
.14
.23
.15
.25
.16
.28
.18
.2
.30
19
.32
.21
.34
.22
.37
.24
.39
.25
.42

.44
.28
.47
.30
.49
.32
.52
-34
.3
.55
.36
.57
.37
.60
.39
.63
.41
.66
.43
.69
.45
.72
.47
.75
.48
.78
.50
.81
52
.4
.84
.54
.87
.56
.91
.59
.94
.61
.97
.63
1.01
.65
1.04
.67
1.07
.69
1.11
.72
1.14
.74
.5
1.18
.76
1.21
.78
1.25
.81
1.28
.83
1.32
.85
1.36
.88
1.40
.90
1.43
92
1.47
.95
1.51
.98
.6
1.55
1.00
1.59
1.03
1.63
1.05
1.67
1.08
1.70
1.10
1.75
1.13
1.79
1.16
1.83
1.18
1.87
1.21
1.91
1.23
.7
1.95
1.26
1.99
1.29
2.03
1.31
2.08
1.34
2.12
1.37
2.16
1.40
2.21
1.43
2.25
1.45
2.29
1.48
2.34
1.51
.8
2.38
1.54
2.43
1.57
2.47
1.60
2.52
1.63
2.56
1.65
2.61
1.69
2.66
1.72
2.70
1.75
2.75
1.78
2.80
1.81
.9
2.84
1.84
2.89
1.87
2.94
1.90
2.99
1.93
3.03
1.96
3.08
1.99
3.13
2.02
3.18
2.06
3.23
2.09
3.28
2.12
1.0
3.33
2.15
3.38
2.18
3.43
2.22
3.48
2.25
3.53
2.28
3.58
2.31
3.63
2.35
3.69
2.38
3.74
2.42
3.79
2.45
1.1
3.84
2.48
3.89
2.51
3.95
2.55
4.00
2.59
4.05
2.62
4.11
2.66
4.16
2.69
4.21
2.72
4.27
2.76
4.32
2.79
1.2
4.38
2.83
4.43
2.86
4.49
2.90
4.54
2.94
4.60
2.97
4.65
3.01
4.71
3.04
4.77
3.08
4.82
3.12
4.88
3 15
1.3
4.94
3.19
4.99
3.23
5.05
3.26
5.11
3.30
5.17
3.34
5.22
3.37
5.28
3.41
5.34
3.45
5.40
3 49
5.46
3.53
1.4
5.52
3.57
5.58
3.61
5.63
3.64
5.69
3.68
5.75
3.72
5.81
3.76
5.87
3.79
5.93
3.84
6.00
3.88
6.06
3.92
IS
6.12
3.96
6.18
3.99
6.24
4.03
6.30
4.07
6.36
4.11
6.43
4.16
6.49
4.19
6.55
4.23
6.61
4.27
6.68
4.32
LEUPOLD & STEVENS, INC
STEVENS WATER RESOURCES DATA BOOK. 2nd ED
-------
EFFLUF.H1 MOM I TOR IMG PROCEDURE: Measurement of Flow in an Open Channel
by Sharp-Crested Weir
Educational Concepts - Mathematics
Section II
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
A float gage is shown in Fig. 3, installed in a
stilling well for measurement of the head on the
weir (H). The floor of the stilling well is level
with the bottom of the channel in which the liquid
is flowing. In order to use the gage to measure
rfT~GAGEINDEX
STILLING WELL
WEIR

FIG. 3 - FLOAT GAGE INSTALLATION.
the head on the weir, it must be "zeroed" under a
set of known conditions, which are, for purposes
of illustration, assumed to be as shown in the
figure. These conditions are as follows:
Height of the weir crest above the channel
floor - 1 foot
Depth of liquid at the gage site - 2 feet
Tape reading opposite gage index - 3 feet
Under these conditions, it is known that the head
on the weir equals one foot, i.e. liquid depth at
the gage (2 feet) minus the distance from the floor
of the channel to the weir crest (1 foot). There-
fore 2 feet must be subtracted from the gage tape
reading to obtain the head on the weir. Conse-
quently, by subtracting 2 feet from the tape
reading under any other condition, the head on the
weir will be obtained.
The following points should be noted in connection
with this procedure:
(a)	If the elevation of the gage index is
changed, the gage must be re-zeroed.
(b)	If the position of the tape on the pulley is
changed, the gage must be re-zeroed.
(c)	The tape must be installed so that the
numerical value of the tape reading In-
creases as the depth of the flow increases.
Page No. 12-15
-------
EFFLUENT MONITORING PROCCDURE: Measurement of Flow in an Open Channel
by Sharp-Crested Weir
Field and Laboratory Equipment
SECTION V
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
A.3
installed is shown in Fig. 4.
LIQUID SURFACE
WEIR CREST
^ NAPPE
HEAD(H)
AIR
FIG. 4 - WEIR INSTALLATION
The following definitions apply:
Weir - A notch of regular form through which
liquid flows.
Weir Crest - The edge over which the liquid
flows.
Sharp-crested Heir - A weir with a sharp upstream
edge so formed that the liquid springs clear of
the crest
Head on Weir (H) - upstream depth of liquid over
the crest of the weir. For a V-notch weir, the
depth is measured from the bottom of the notch.
Nappe - the overflowing sheet of liquid.
Free Discharge (free-flow) - when nappe
discharges into the air.
Submerged Discharge (submergence) - when liquid
level aownstream of the weir is at a higher
elevation, or the same elevation as the weir
crest, so that the nappe discharges partially
under water.
Zero Gacie Elevation - The division on the staff
gage which is at the same level as the weir
crest.
A.4
Weirs are designated according to the shape of the
notch through which the liquid flows. The types
of weirs most commonly used to measure wastewater
flows are:
Paqe No. 12-16
-------
EFFLUENT MONITORING PROCEDURE: Measurement of Flow in an Open Channel
by Sharp-Crested Weir
Field and Laboratory Equipment
Section V
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
A.4 (Cont'd)
1. The 50° V-notch (triangular) weir, which has
sides inclined 45° from the vertical.
METAL
METAL
PLATE
SECT. A-A
FIG. 5 - 90° V-NOTCH WEIR
(NOTE: Triangular weirs having notch angles other
than 90° may also be used. Angles of 22-1/2°,
45°, and 60°, will sometimes be seen. Procedures
for using these weirs are exactly the same as for
the 90° weir, except that different formulas and
weir tables apply. It is necessary that the
proper formula or_£able be selected for the specific
weir being used.)
2. The standard contracted rectangular weir, or
weir with end contractions.
L—»
CREST
LA
§
L=CREST
4- LENGTH
SEE FIG. 5
FOR SECT. A-A
FIG. 6 - WEIR WITH END CONTRACTIONS
Paqe No. 12-17
-------
EFFLUi-fn HON I TORI MO PROCEDURE: Measurement of Flow in an Open Channel
by Sharp-Crested Weir
Field and Laboratory Equipment
Section V
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
A.4 (Cont'd;
3. The standard suppressed rectangular weir, or
weir without end contractions.
L=CREST
LENGTH
SEE FIG. 5
FOR SECT. A-A
A.5
FIG. 7-SUPPRESSED WEIR
The weir notch or weir crest, as shown in the above
illustrations, is cut with a sharp upstream edge
into a relatively thin metal plate that is mounted
on a supporting bulkhead. The crest should be 1-2 mr
thick, (3/64 to 5/64 inch).
A standard staff gage, used for obtaining head
measurements, is illustrated below:
FIG. 8 - STAFF
GAGE SECTION
Commercially-available gages are generally made of
18 gage metal coated with a substantial thickness of
porcelain enamel. The standard gage is 4" wide and
3-1/3' long. The face of the gage is white; numerals
and graduations are black. Gages may be made to any
length desired, using similar details.
A float gage (Fig. 9) is a means of continuously
indicating liquid levels. It consists of a metal
float, a pulley mounted on a standard, and a
Page No. 12-18
-------
EFFLUENT MONITORING PROCEDURE: Measurement of Flow in an Open Channel
by Sharp-Crested Weir
Field and Laboratory Equipment
Section V
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
A.6 (Cont'd^
FIG. 9 - FLOAT GAGE
counterweight. A graduated stainless steel tape is
attached to the float and connected at the other
end to the counterweight. The float follows the
rise and fall of the liquid surface and the level
can be read from the tape and a pointer or
reference mark. Tapes are available in selected
lengths, and are graduated either in feet, tenths,
and hundredths for English measurements, or meters,
decimeters and centimeters for metric measurements.
The float gage is used extensively as a reference
gage 1n stilling wells to check the accuracy of
automatic head or flow recording devices.
The measured head will, be too low If leakage of the
liquid occurs along the sides or bottom of the
bulkhead. All observed leaks should be immediately
elIminated.
The upstream face of the bulkhead should be 1n a
vertical plane perpendicular to the axis of the
channel, for accurate results.
The bulkhead should be perpendicular to the
direction of liquid flow, for accurate results.
The weir crest must be horizontal for standard
formulas and weir tables to apply. The crest
should be checked periodically, and leveled if
required.
Page No. 12-19
-------
EFFLUENT MONITORING PROCEDURE: Measurement of Flow in art Open Channel
by Sharp-Crested Weir
Field and Laboratory Equipment
Section V
TRAINING GUIDE MOTE
REFERENCES/RESOURCES
The gage division which is at the elevation of the
weir crest will be referred to as the "zero gage
elevation." Its value must be known in order to
calculate the head on the weir. When a weir in-
stallation is made, the zero gage elevation is
determined, but, since this may change for some
reason, it should be checked from time to time and
a new zero elevation established, if necessary.
Small nicks and dents can reduce the accuracy of a
weir installation. Those that do occur should be
carefully dressed with a fine-cut file or stone,
stroking only in the plane of the upstream weir
face, the plane of the weir crest or sides, or the
plane of the chamfers. Under no circumstances
should the upstream corners of the notch be rounded
or chamfered; nor should the shape of the weir
opening be changed by attempting to completely re-
move an imperfection. Instead, only those portions
of the metal that protrude above the normal surfaces
should be removed. In extreme cases, replacement
of the weir plate may be required.
Build up of extraneous material on the weir crest
can cause inaccurate results. Such material should
be cleaned off the weir plate prior to a head
measurement.
If the liquid level downstream of the weir rises
high enough so that there 1s no air space under the
nappe, use of standard formulas and weir tables will
produce inaccurate results. The nappe must be
ventilated, i.e., have an air space underneath it.
Do not attempt to use the weir as a measuring device
if it is operating under a condition of submerged
di scharge.
If the nappe does not spring completely free of the
weir, but clings to the downstream side wholly or 1n
part, an inaccurate result will be obtained. The
cause of such a condition must be determined, and
the condition corrected, if good data are to be
secured.
Any large submerged or floating objects in the
channel upstream of the weir should be removed.
Sediment deposits behind the weir structure can
affect the accuracy of the installation. Deposited
material must be cleaned out when the vertical dis-
tance from the top of the deposit to the weir crest
Page No. 12-20
-------
EFFLUENT MONITORING PROCEDURE: Measurement of Flow in an Open Channel
by Sharp-Crested Weir
Field and Laboratory Equipment		Section V
TRAINING GUIDE MOTE
REFERENCES/RESOURCES

is one foot or less. More frequent cleaning is
desirable.

B.1.5
A disturbance of the normal flow pattern will affect
the accuracy of a measurement made while such dis-
turbance exists. If the normal flow is disturbed
for any reason in connection with obtaining a head
reading, adequate time should be allowed before
making the reading, so that normal conditions may
be re-established.

Page No. 12-21
-------
A PROTOTYPE FOR DEVELOPMENT OF
ROUTINE OPERATIONAL PROCEDURES
for the
AMPEROMETRIC DETERMINATION OF FREE AND COMBINED
RESIDUAL CHLORINE IN WATER
as applied in
WASTEWATER TREATMENT FACILITIES
and 1n the
MONITORING OF EFFLUENT WASTEWATERS
Developed by the
National Training and Operational Technology Center
Municipal Operations and Training Division
Office of Water Program Operations
U.S. Environmental Protection Agency
CH.CL.EMP.2a.3.77
Page No. 13-1
-------
EFFLUENT MONITORING PROCEDURE: Arnperometric Determination of Free and Combined
Residual Chlorine in Water
This instructional sequence was developed by:
NAME Paul F. Hall bach
ADDRESS EPA, OUPO. NTOTC. Cincinnati. Ohio
POSITION Chemist Instructor
EDUCATION AND TECHNICAL BACKGROUND
B.S. - Chemistry
14 years Industrial Chemist
16 years HEW-FWPCA-EPA-Chemist
Page No. 13-3
-------
EFFLUENT MONITORING PROCEDURE: Amperometric Determination of Free and Combined
Residual Chlorine in Water
1.	Analysis Objectives
The operator will be able to perform an amperometric titration for the determination
of free and combined residual chlorine in water.
2.	Brief Description of Analysis*
Free available residual chlorine and combined residual chlorine are titrated
successively using an amperometric titrator. The free available residual chlorine
is titrated first. The sample pH is then dropped to 4 by adding buffer solution
pH 4 and then potassium iodide is added to the sample. The first titration will
represent the free available residual chlorine while the second titration will
represent the combined residual chlorine.
3.	Applicability of this Procedure:
a.	Range of Concentration:
Chlorine residuals over 2 mg/1 are best measured by means of smaller samples,
or by dilution with water that neither is chlorinated nor has a chlorine demand.
b.	Pretreatment of Samples:
None
c.	Treatment of Interferences in Samples:
None
~Standard Methods for the Examination of Water and Wastewater, 14th Ed., 1975. APHA.
Washington, D.C., p. 322
Page No. 13-4
-------
EFFLUENT MONITORING PROCEDURE: Amperometric Determination of Free and ComM'ned
Residual Chlorine in Water
General Description of Equipment used in the Process
A. Capital Equipment
1. Amperometric Titrator Assembly - Wallace and Tiernan*
G. Reusable
1.	1	pipette {1 nl capacity)
2.	1	pipette (5 ml capacity)
3.	1	sample cup (to contain 200 ml)
4.	1	plastic squeeze bottle
C. Consumable**
1.	1	bottle phenylarsene oxide solution 0.00564 N (16 ounce)
2.	1	bottle pH 4 buffer solution (4 ounce)
3.	1	bottle pH 7 buffer solution (4 ounce)
4.	1	bottle potassium iodide solution (4 ounce)
5.	1	bottle sodium chloride electrolyte tablets (8 ounce)
~Mention of a specific brand name does not constitute endorsement by the
U.S. Environmental Protection Agency
~~Consumable reagents listed are available from Wallace & Tlernan Industrial
Products Division, 25 Main St., Belleville, Nd 07109
Page No. 13-5
-------
EFFLUENT MONITORING PROCEDURE: Amperometric Determination of Free and
Combined Residual Chlorine in Water
Paae No. 13-6
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
A. Equipment Preparation
1.
Set up titrator on work
bench.
la. Electric outlet 110 volt required.
lb. Amperometric titrator assembly available from
Wallace and Tiernan Corporation


2.
Select proper pipette for
titration.
2a. Two pipettes are furnished with the titrator.
The 1 ml pipette is generally used when the
residual is less than 1 mg/1. A 5 ml pipette is
for use with higher residuals.


3.
Lightly grease the lower
end and insert it in the
top of the pump unit on the
side of the titrator.
3a. Use silicone grease or other similar lubricant.


4.
Fill the pump squeeze bottle
about 2/3 to 3/4 full with
phenylarsene oxide solution.
4a. Reagent is highly toxic - avoid ingestion.


5.
Screw the bottle on to the
pump.
5a. It is easier to turn the bottle than the cap.


6.
Pour sufficient electrolyte
tablets into the cell unit
to fill the chamber about
2/3 full.

VII.A.6
(p. 15)

7.
Add enough distilled water
to cover the tablets.
7a. Use a plastic squeeze bottle.


8.
Plug the cell unit into the
titrator.
8a. The cell is so designed that it cannot be plugged
in except in the correct position.
V.A.8.8a
(p. 13)

9.
Examine the titrator cup.
The cup has a line indicat-
ing the 200 ml level.
9a. Whenever the term "sample" is used in these in-
structions it shall mean a 200 ml volume of the
water to be tested.
1
-------
EFFLUENT MONITORING PROCEDURE: Amperometric Determination of Free anc!
—— ~	Combined Residual Chlorine in Water
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
B. Determination of Free
Available Residual
Chlorine
1.	Plug the electric power plug
into a source of 115 volt,
single phase, 60 cycle A.C.
current.
2.	Fill the pipette with
phenylarsene oxide solution.
3.	Remove all air from the
pipette and plastic tubing
by rotating the red knob in
the stem unit 1/4 turn
counter-clockwi se.
4.	Catch the discarded solution
in a 50 ml beaker.
5.	Refill the pipette to the
top (zero) calibration mark.
6.	Add sample water to the cup.
Adjust the level to the line.
7.	Place the cup on the
titrator.
2a. Alternately squeeze and release the squeeze
bottle.
3a. The pipette should drain through the plastic
tubing.
6a. The volume of sample is 200 ml.
7a. The top edge of the cup should go behind the cup
guide post.
7b. The bottom of the cup should rest on the support
post.
7c. The plastic tubing from the pump should be sub-
merged in the sample about 1/16 inch. If
necessary, adjust the tubing on the guide post
to obtain this condition.
I.B.I
{p. 13)
Page No. 13-7
-------
EFFLUENT MONITORING PROCEDURE: Amperometric Determination of Free and
Combined Residual Chlorine in Water
Page No. 13-8
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
Add 1 ml of buffer solution
pH 7 to the water sample.
9. Start the agitator by turn-
ing the switch to "ON".
JC. Adjust the meter to make the
pointer read maximum on the
scale.
11. Start adding small amounts
of titrant and note the
deflection of the meter
scale after each addition.
12. Continue the addition of
small amounts of titrant un-
til the addition of titrant
no longer causes a deflection
of the needle.
8a. The droppers furnished with the titrator are 1 ml
units. A dropper full of solution should be used
wherever 1 ml of solution is called for.
8b. If the pH of the sample is between 6.0 and 7.5
it is not necessary to add buffer.
10a. Rotating the adjusting knob clockwise should
increase the reading.
10b. If the pointer is above maximum when the adjust-
ing knob is rotated completely counter-clockwise,
then the titration should be started with the
knob in this position.
11a. If free available chlorine is present in the
sample and if the pointer is on scale at the
beginning of the titration, then the first addi-
tion of titrant should cause a definite pointer
movement to the left. If the pointer goes below
zero then it should be brought back on scale by
rotating the adjusting knob clockwise.
12a. In most waters the end-point of the reaction is
just passed when the addition of a small amount
of titrant no longer deflects the pointer to the
left.
12b. The amount of titrant used in the titration is
then read fron the pipette and the last increment
is subtracted from the pipette reading and the
resultant figure represents the free available
residual chlorine in mg/1.
-------
EFFLUENT MONITORING PROCEDURE: Amperometric Determination of Free and
Combined Residual Chlorine in Water
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES

^3. Subtract the last reading
from the previous reading.
14.	The reading on the pipette
represents the amount of
free available chlorine in
mg/1.
15.	Turn instrument "OFF".
16.	Record your result.


C. Determination of
combined residual
chlorine
1.	Repeat steps 1 through 7 of
the free available chlorine
procedure if the free avail-
able chlorine determination
has not been performed.
2.	If you have just completed
the free chlorine determina-
tion, you can continue the
use of the same sample for
this determination.
3.	Add 1 ml of buffer solution
pH 4 to the sample.
4.	Add 1 ml of potassium iodide
solution to the water sample.
la. The general procedure for measuring total residual
chlorine is the same as that given for measuring
free available residual chlorine.
3a. Use the dropper to add the buffer solution.
4a. Use the dropper to add the potassium iodide
solution.
4b. When potassium iodide is added, the pointer may
first deflect to the left and then go up-scale.
VII.C.4.4b
(P. 14)
Page No. 13-9
-------
EFFLUENT MONITORING PROCEDURE: Amperometric Determination of Free and
Combined Residual Chlorine in Water
Page No. 13-10
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
trai:; i :;g
GUIDE .NOTES

5. Follow steps 9 through 16 of
the previous procedure for
the determination of free
available chlorine. In this
case the result is reported
as combined residual
chlorine.
5a. Free available residual chlorine and combined
residual chlorine may be measured in one sample
by combining the two procedures.
5b. The free available chlorine is measured first.
The sample pH is then dropped to 4 by adding
buffer solution pH 4 and then potassium iodide.
5c. If combined residual chlorine is present, the
pointer will deflect to the right when potassium
iodide is added.
5d. The first titration will represent the free
available residual chlorine while the second
titration will represent the combined residual
chlorine.

-------
E F F L U ENT MOM IT0RING PROCEDURE:
Amperometric Determination of Free and
Combined Residual Chlorine in Water
Sections I & V
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
I.B.I
V.A.8.8a
The fundamental chemical procedure involved
in the amperometric titrator is the neu-
tralization of an oxidizing agent (free
available chlorine) in a sample of water by
the addition of a reducing agent of known
strength. Immersed in the sample cell unit
which produces a small direct current which
is proportional to the free chlorine present
in the sample. The current is indicated on
a microammeter which is connected to the
cell unit. As the reducing agent is added,
the amount of free chlorine is reduced, the
cell current decreases, and the micro-
ammeter pointer moves down scale. The end
point of the reaction occurs when enough
reducing agent has been added to just
neutralize all of the free chlorine in the
sample. When this point is reached, the
further addition of a small amount of re-
ducing agent no longer deflects the pointer
to the left. On the titrator, the sample
volume and the strength of the reducing
agent have been selected to make 1 milli-
liter of reducing agent equivalent to one
milligram per liter of chlorine. When the
endpoint is reached, therefore, the volume
of reducing agent used represents the
chlorine concentration in mg/1.
Under the conditions specified in the
titration procedure, the titration can be
used to distinguish between free available
residual chlorine and combined residual
chlorine because the reducing agent em-
ployed reacts readily with free chlorine
but does not react with combined chlorine.
If either combined or total residual
chlorine is to be measured, potassium
iodide is added to the sample to produce
an amount of free iodine which is equiva-
lent to the original residual chlorine.
The reducing agent reacts readily with free
iodine so that the titration can be carried
out in a manner similar to that used for
free available residual chlorine
determination.
The electrolyte used in the inner chamber of
the cell has a tendency to crystallize out
on the contact springs and in the terminals
of the cell unit. This may slightly corrode
Page No. 13-12
-------
EFFLUENT MONITORING PROCEDURE: Amperometric Determination of Free and
Combined Residual Chlorine in Water
TRAINING GUIDE
SECTION	TOPIC
I*	Introduction
II	Educational Concepts - Mathematics
III	Educational Concepts - Science
IV	Educational Concepts - Communications
V*	Field and Laboratory Equipment
VI	Field and Laboratory Reagents
VII*	Field and Laboratory Analysis
VIII	Safety
IX	Records and Reports
~Training guide materials are presented here under the heading marked *.
These standardized headings are used throughout this series of procedures.
Paqe No. 13-11
-------
EFFLUENT MONITORING PROCEDURE: Amperometric Determination of Free and
Combined Residual Chlorine in Water
Sections V & VII
TRAINING GUIDE MOTE
REFERENCES/RESOURCES
VII.C.4.4b
the electrical contacts between the various
units. Improper electrical connections
cause erratic microammeter pointer readings
during the titration. Should any crystals
accumulate on the plastic cell unit, these
parts should be washed off with warm water.
CAUTION: Never use water warmer than
100°F, as hot water softens the plastic.
When the titrator is not to be used for
extended periods, the cell unit should be
washed out to remove all electrolyte
tablets and solution, and stored dry.
If free available residual chlorine de-
terminations are to be made after potassium
iodide has been used in preceding titrations,
the cell unit should be rinsed off in
several sample cups of water to remove
traces of potassium iodide solution and
buffer solution pH 4.
Occasionally, when potassium iodide is added]
to the sample, the pointer will drop to the
left and will not come back on scale even
though the poteniometer is turned com-
pletely clockwise. Under these conditions,
the cell unit is said to have lost its
sensitivity to iodine. This situation is
likely to arise if the titrator has been
used to determine free chlorine only for
extended periods of time, i.e., the cell
unit has not been exposed to iodine for
prolonged periods.
The sensitivity of the cell unit can be re-
stored by adding enough free iodine to the
distilled water in the sample jar to create
a yellowish color. The free iodine may be
in the form of tincture of iodine or may be
obtained by adding potassium iodide to a
strong chlorine solution. Agitate the
sample for two or three minutes and then
allow the cell unit to stand in the iodine
solution for 10 to 15 minutes. After this
treatment, the cell unit should be rinsed
off throughly to remove all traces of
iodine.
Page No. 13-13
-------
EFFLUENT f-'fitiITORING PROCEDURE: Amperometric Determination of Free and
"	Combined Residual Chlorine in Water
Section VII
TRAINING GUIDE MOTE
REFERENCES/RESOURCES
VII.A.6
The main requirement as far as electrolyte
tablets are concerned is to have saturated
electrolyte solution inside the cell unit at
all times. Theoretically, this requirement
is not as long as any tablets and water are
in the cell unit. The actual water level
inside the cell unit cannot be controlled
since this level tends to equalize with
(or even go below) the water level in the
sample jar through the porous wicking.

Page No. 13-14
-------
A PROTOTYPE FOR DEVELOPMENT OF
ROUTINE OPERATIONAL PROCEDURES
for the
AMPEROMETRIC DETERMINATION OF TOTAL
RESIDUAL CHLORINE IN WASTEWATER
as applied 1n
WASTEWATER TREATMENT FACILITIES
and 1n the
MONITORING OF EFFLUENT WASTEWATERS
Developed by the
National Training and Operational Technology Center
Municipal Operations and Training Division
Office of Water Program Operations
U.S. Environmental Protection Aaency
CH.CL.EMP.3.3.77
Page No. 14-1
-------
EFFLUENT MONITORING PROCEDURE: Amperometric Determination of Total Residual
Chlorine in Wastewater
This instructional sequence was developed by:
NAME Paul F. Mall bach
ADDRESS EPA, OIJPO* NTOTC, Cincinnati, Ohio 45268
POSITION Chemist-Instructor
EDUCATION AND TECHNICAL BACKGROUND
B.S. - Chemistry
14 years Industrial Chemist
16 years HEW-FWPCA-EPA-Chemi st
Page Mo. 14-3
-------
EFFLUENT MONITORING PROCEDURE: Amperometric Determination of Total Residual
Chlorine in Wastewater
1.	Analysis Objectives
The operator will be able to perform an amperometric titration for the determination
of total residual chlorine in a sample of wastewater treatment plant effluent.
2.	Brief Description of Analysis*
Residual chlorine present in wastewater is in the	form of combined chlorine.
A "Back-Titration" procedure is used to determine	the phenylarsene oxide excess
and a formula used to calculate the concentration	of total residual chlorine in
the sample.
3.	Applicability of this Procedure:
a.	Range of concentrations:
Applicable to all types of wastewater.
b.	Pretreatment of Sample
None
c.	Treatment of Interference 1n Samples.
Manganic Manganese, in concentrations as low as l.Omg/liter liberates iodine
from iodide at a pH of 4.0.
Chromates reduce phenylarsine oxide. Method not applicable when high concentra-
tions of chromates are present.
Annual Book of ASTM Standards, 1975. American Society for Testing and Materials.
1916 Race St., Philadelphia, PA 19103. p. 278.
Page No. 14-4
-------
EFFLUENT MONITORING PROCEDURE: Amperometric Determination of Total
Residual Chlorine in Wastewater
General Description of Equipment used in the Process
A.	Capital Equipment
1. Amperometric Titrator Assembly - Wallace and Tiernan*
B.	Reusable
1.	1	pipette (1 ml capacity)
2.	1	pipette (5 ml capacity)
3.	1	sample cup (to contain 200 ml)
4.	1	plastic squeeze bottle
C.	Consumable**
1.	1 bottle phenylarsene oxide solution 0.00564N (16 ounce)
2.	1 bottle pH 4 buffer solution (4 ounce)
3.	1 bottle pH 7 buffer solution (4 ounce)
4.	1 bottle potassium iodide solution (4 ounce)
5.	1 bottle sodium chloride electrolyte tablets (8 ounce)
6.	Standard Iodine solution 0.1 N
7.	Standard iodine titrant 0.0285"^
8.	Potassium iodide crystals
9.	Iodine crystals, purified
~Mention of a specific brand name does not constitute endorsement by the
U.S. Environmental Protection Agency
~~Consumable reagents listed are available from Wallace * Hernan Industrial
Products Division, 25 Main St., Belleville, NJ 07109
Page No. 14-5
-------
EFFLUENT MONITORING PROCEDURE: Amperometric Determination of Total Residual Chlorine in Wastewater
Page No. 14-6
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
A. Reagents
Standard iodine
solution 0.1 N
Iodine standard
solution (0.0282H)
1.	Dissolve 40.0 grams of
potassium iodide (KI) in
50 ml of distilled water.
Add 12.7 g. of iodine
crystals and stir until
solution is complete.
2.	Dilute to one liter with
distilled water.
3.	Transfer 25 grams of
potassium iodide into a
one liter volumetric flask.
4.	Add 200 ml of distilled
water and swirl to dissolve.
5.	Add 285 ml of 0.1 N iodine
solution and dilute to the
mark with distilled water.
2a. Store the solution in a dark bottle.
3a. Use a trip balance.
4a. Use a graduate cylinder.

B. Determination of
total residual
chlorine
1.	Set up titrator and plug
into a source of 115 volt,
single phase, 60 cycle A.C.
current.
2.	Add sample water to the cup.
Adjust the level to the
1 i ne.
3.	Place the cup on the
titrator.
2a. The volume of sample is 200 ml.
3a. The top edge of the cup should go behind the cup
guide post.
3b. The bottom of the cup should rest on the support
post.

-------
EFFLUENT MONITORING PROCEDURE: Amperometric Determination of Total Residual Chlorine in Wastewater
DERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
Continued
4.	Turn the switch to start
the agitator.
5.	Add 5 ml of phenylarsene
oxide solution to the sample
and mix.
6.	Add 4.0 ml of pH 4.0 buffer
solution to the sample and
mix.
7.	Add 1.0 ml of KI solution.
8.	Rotate the adjusting knob so
that the microarmeter pointer
reads about 20 on the scale.
9.	Add 0.0282 iodine solution
in small Increments.
0. Note the volume of iodine
solution used to reach the
end-point.
5a. Use a 5 ml pipette.
6a. Should be sufficient to insure a sample pK
between 3.5 and 4.2.
9a. Use a 1 ml volumetric pipette graduated in 0.1 ml.
9b. The standard reagent bottle, pump, pipette, and
applicator tubing cannot be used for this DurDose
since the plastic components may react with the
iodine solution and change its strength.
9c. As iodine is added to the sample, the pointer re-
mains practically stationary until the end-point
is approached. Just before the true end-point
each increment of iodine solution causes a
temporary deflection of the microaimieter to the
right, but the pointer drops back to about its
original position. The true end-point is reached
when a small addition of iodine solution gives a
definite and permanent pointer deflection to the
right (up-scale).
Oa. Calculate the total residual chlorine as follows:
total ] (5) (ml of iodine)"
mg/1 chlorine = phenylarsenef - used in
oxide used . titration
(step 5) J

Page No. 14-7
-------
EFFLUENT MONITORING PROCEDURE: Amperometric Determination of Total Residual Chlorine in Wastewater	Page No. 14-8
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
E. Continued

10b. Example of calculation:
1.	Total phenylarsene oxide used in
step 5 = 5.0 ml.
2.	ml of iodine required to reach the end-point
in step 9 = 0.6 ml
mg/1 chlorine = 5.0 - (5)(0.6)
= 5.0 - 3.0
= 2.0
10c. The accuracy of the above procedure depends on the
volume of the sample (step 2), ttie strength of
the phenylarsene oxide solution (0.00564N) which
is quite stable, and the strength of the iodine
solution (0.0282 N) which is subject to
deterioration with time. If the iodine is not
0.0282 N, it must be standardized by the following
procedure.

C. Standardization of
iodine solution *
!• Add 5,0 ml of phenylarsene
oxide solution to 195 ml of
dechlorinated water.
2. Titrate with the iodine
solution.
la. Chlorine-demand-free water: add sufficient
chlorine to distilled water to destroy the
ammonia. The amount of chlorine required will be
about ten times the amount of ammonia nitrogen
present; in no case produce an initial residual
of less than 1.0 mg/1 free chlorine. Allow the
chlorinated water to stand overnight or longer;
then expose to direct sunlight until all residual
chlorine is discharged. Use distilled water free
from ammonia and nitrite to produce the chlorine
demand-free water. Check chlorine residual by
amperometric titration.
2a. The end point is reached when a small addition of
iodine gives a pointer deflection to the right
(up scale) which holds for 15 to 20 seconds. If
1.0 ml of iodine solution neutralizes the 5.0 ml
of phenylarsene oxide solution, the iodine solution

-------
EFFLUENT MONITORING PROCEDURE: Amperometric Determination of Total Residual Chlorine in Wastewater
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES

12.	Locate on line "A" the ml
of iodine equal to 5.0 ml
of phenylarsene oxide.
13.	Locate on line "C" the
volume of iodine as de-
termined in step 9.
14.	Determine where a line
connecting these points
crosses line "B".
11a. Continued
is 0.0282 N. If the iodine solution has
deteriorated, the volume of iodine solution to
reach the end-point (something greater than
1.0 ml) is equal to 5 ml of phenylersene oxide
solution.
lib. "Back titration" for residual chlorine may be
made with weaker than 0.0282 N iodine solutions.
The attached chart can be used to determine the
excess phenylarsene oxide by following step 12
and subsequent steps.
14a. This is the excess phenylarsene oxide.
14b. As expressed in the formula, the mg/1 of chlorine
residual is the excess phenylarsene oxide sub-
tracted from the total.
14c. Example of calculation:
1.	Total phenylarsene oxide ® 10.0 ml
2.	ml iodine equal to 5.0 ml phenylarsene
oxide = 1.2 ml.
3.	ml iodine to reach end point of "back
titration" = 0.4
4.	Excess phenylarsene oxide (from chart) = 1.6
(approx.)
5.	mg/1 chlorine residual = 10-1.6 = 8.4

Page No. 14-9
-------
EXAMPLE
Total phoriylarsene oxidiv-5 ml.
ml. iodine equal to 5 ml. phenylarseria oxide	-1 ml.	(0.0282N solution)
ml. iodine to reach end point of "Back Titration" -	0.6
Excess phenylarsene oxide (from chart)	~	3.0
or (from formula)	=	5 x 0.6 3.0
ppm chlorine; residual	~	5 - 3 ~2
EXAMPLE
Total plv^nyluriona or.ide --I0 ml.
ml. iodir.e equal to 5 ml. phsnyia. renc: oxide -1.2 ml.
ml. iodine to rec;ch end point of "Bock Titration"
Excess pht-iiylarseno oxide (from chart)	-1.6 (approximately)
ppm chlorine residual	"10- 1.6-8.4
CO
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U!
X
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I-
<£
.o
a
w
Ui
o
u.
o
E
Jjl.0 (0.0282N)
\
\
\
\
\
-I.I
\
\
V
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s
b
c
/:
i-.
t
[:
»-
:!.6
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V
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S. ^
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\
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\
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20
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-10
-7.5
UI
e u.o
\ X
\o L-,0
\
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. UI
V)
ITt
'ji
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in
i»i
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2.5.
2.0 >0.6
0.5
'-1.5
1.1>
I. 9
2.0
1.0
-0.75
0.!>
•0.25
0.4
0.3
0.2
0.15
0.1
Page No. 14-104 *
e>
z
o
9.0
7.0
-6.0
-5.0
'.0
-o.O
-2.0
-1.5
1.0
-0.9
-0.8
0.7
-------
A PROTOTYPE FOR DEVELOPMENT OF
ROUTINE OPERATIONAL PROCEDURES
for the
TITRIMETR1C DETERMINATION OF TOTAL
RESIDUAL CHLORINE IN WASTEWATER EFFLUENTS
as applied in
WASTEWATER TREATMENT FACILITIES
AND IN THE
MONITORING OF EFFLUENT WASTEWATERS
Developed by the
National Training and Operational Technology Center
Municipal Operations and Training Division
Office of Water Program Operations
U.S. Environmental Protection Agency
CH.CL.EMP.4.6.77
Page No.
15-1
-------
EFFLUENT MONITORING PROCEDURE: Titrimetric Determination of Total Residual
Chlorine in Wastewater
This operational procedure was developed by:
NAME	Charles R. Feldmann
ADDRESS EPA, OWPO, NTOTC, Cincinnati, Ohio 45268
POSITION Chemist-Instructor
EDUCATION AND TECHNICAL BACKGROUND
B.S. - Chemistry
M.S. - Chemistry
1-1/2 years Industrial Chemist
4 years additional Graduate School
4 years college Chemistry Instructor
1-1/2 years DHEW - Air Pollution Program, Chemist
10 years 01 - EPA, Chemist-Instructor
Page No. lb-J
-------
EFFLUENT MONITORING PROCEDURE: Titrimetric Determination of Total Residual
Chlorine in Wastewater Effluents
1.	Analysis Objectives:
The learner will determine the total residual chlorine in a sample of
wastewater treatment plant effluent.
2.	Brief Description of Analysis:
Chlorine, hypochlorus acid, and hypochlorite ion are collectively referred
to as free chlorine. Free chlorine is added to wastewater effluents for
disinfection purposes. It combines with ammonia in the effluent to form
monochloramine, dichloramine, and nitrogen trichloride; these three com-
pounds together, are called combined chlorine. The sum of combined and
free chlorine is referred to as total residual chlorine. In this procedure,
called a back-titration, an amount of reducing agent (phenylarsene oxide)
more than sufficient to react with the total residual chlorine is added to
the sample. The amount of excess reducing agent is then determined by
titration with standard iodine solution. The result is expressed as mg of
total residual chlorine per liter of sample.
3.	Applicability of this Procedure:
a.	Range of Concentration:
Although the cited reference* does not specifically mention the range
of applicability, it can be inferred from the procedure that concentra-
tions of up to 10 mg total residual chlorine/liter can be accurately
analyzed. No inference can be made about the lower limit of the test.
b.	Sample Pretreatment:
None. The determination must be carried out immediately after sampling.
Avoid exposure of the sample to strong sunlight and excessive agitation.
c.	Treatment of Interferences:
The interference due to manganese, iron, and nitrite is minimized by
buffering the reaction mixture as described in this procedure. If the
sample contains a large amount of organic matter, the titration end-
point may be obscured. This problem may be overcome by acidifying the
reaction mixture to a pH of 1.0, but only if the sample contains no
manganese, iron, or nitrite. If they are present, then the amperometric
procedure should be used.
* Source of Procedure: Standard Methods, 14th ed., method 409 B., page 318.
Page No. 15-4
-------
EFFLUENT MONITORING PROCEDURE: Titrimetric Determination of Total Residual
Chlorine in Wastewater Effluents
General Description of Equipment Used in the Process
A.	Capital Equipment:
1.	Trip balance, 100 g capacity
2.	Analytical balance
3.	Still, or other source of distilled water
B.	Reusable Supplies:
1.	Brushes (for cleaning glassware)
2.	Brush (for cleaning balance)
3.	Laboratory apron
4.	Safety glasses
5.	One distilled water plastic squeeze bottle
6.	One pen or pencil
7.	One notebook (for recording data)
8.	Sponge (for cleaning laboratory table top)
9.	One 2 liter Erlenmeyer flask
10.	One 500 ml Erlenmeyer flask
11.	One 250 ml Erlenmeyer flask
12.	One 125 ml Erlenmeyer flask
13.	One 1 liter graduated cylinder
14.	One 500 ml graduated cylinder
15.	One 250 ml graduated cylinder
16.	One 100 ml graduated cylinder
17.	One 10 ml graduated cylinder
18.	Six 1 liter glass-stoppered bottles
19.	One 1 liter plastic bottle
20.	Two 100 ml glass-stoppered bottles
21.	One 1 liter volumetric flask
22.	One 50 ml volumetric pi pet
23.	One 10 ml volumetric pipet
24.	One 5 ml volumetric pipet
25.	One 10 ml graduated pipet
26.	One 250 ml beaker
27.	One 30 ml beaker
28.	One 2 liter beaker (for cleaning glassware)
29.	One small spatula (for use when weighing solids)
30.	One hot plate (to accommodate a 2 liter Erlenmeyer flask)
31.	One mortar and pestle (about 100 ml capacity)
32.	One 50 ml buret
33.	One 5 ml buret
34.	One small powder funnel (to fit into the top of a 1 liter volumetric flask)
35.	One small funnel (to fit in the top of the buret)
36.	One clamp (to support the buret)
37.	One ring stand (for use with the buret and clamp)
38.	Magnetic stirrer and 2 inch stirring bar (optional)
39.	One weighing bottle with top (about 15 ml capacity)
40.	Fifteen inches of 6 mm glass tubing
41.	Two feet of tygon tubing (to connect the lecture bottle of carbon
dioxide to the 6 mm glass tubing)
Page No. 15-5
-------
EFFLUENT MONITORING PROCEDURE: Titrimetric Determination of Total Residual
Chlorine in Wastewater Effluents
42.	One	universal clamp (to support the lecture bottle on the ring stand)
43.	One	eyedropper
44.	One grease pencil
45.	One	pH meter (with pH 4 & 7 buffers)
46.	Six	inch stirring rod
47.	Sufficient aluminum foil to wrap a 1 liter glass-stoppered bottle
48.	One	asbestos glove or towel (to facilitate lifting a flask of hot water)
C. Consumable Supplies:
1.	Concentrated sulfuric acid, H^SO^
2.	Sodium dichromate, Na2Cr207
3.	Soap
4.	Eight plastic weighing boats (about 2 inches square)
5.	76 g of potassium iodide, KI
6.	5 g of soluble starch
7.	1.25 g salicylic acid, 2-H0CgH4C02H
8.	5 g of arsenic trioxide, As^
9.	27 g of sodium hydroxide, NaOH
10.	Lecture bottle of carbon dioxide, COg
11.	13 g of resublimed (some catalogs simply use the word sublimed) iodine, I«
12.	55 ml of concentrated (12N) hydrochloric acid, HC1
13.	0.8 g of phenyl arsine oxide powder, CgHgAsO
14.	146 g of anhydrous sodium acetate, NaCjH-A, or 243 g of sodium acetate
trihydrate, NaC2H302*3H20
Items C.I., C.2., and C.3. are for cleaning glassware. The quantities needed
will therefore vary.
Items C.5. through C.14 (except C.10.)
weighing solids and measuring liquids,
amounts.
are the exact amounts needed. To facilitate
include slightly more than the required
Page No. 15-6
-------
EFFLUENT MONITORING PROCEDURE: Titrimetric Determination of Total Residual
Chlorine in Wastewater Effluents
All reagents should be of high quality. Different chemical manufacturers may
have different ways of indicating a high quality reagent. While no endorsement
of one chemical manufacturer over another is intended, the following are some
designations used in four chemical catalogs to indicate high quality reagents.
Catalog
Thomas
Matheson, Coleman & Bell
Curtin Matheson Scientific, Inc.
Fisher
Designations
Reagent, ACS, Chemically Pure (CP)
Reagent, ACS
Primary Standard, ACS, AR
Certified, ACS
Page No. 15-7
-------
Page No. 15-8
RING STAND
LECTURE
BOTTLE OF
CARBON
DIOXIDE
UNIVERSAL
CLAMP
-------
EFFLUENT MONITORING PROCEDURE: Titrimetric Determination of Total Residual Chlorine in Wastewater Effluents
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
A. Equipment Preparation
1.	Cleaning of glass-
ware
2.	Balance inspection
1. Clean all glassware and
rinse with distilled water.
1. Check the analytical and
trip balances for cleanli-
ness and proper operation.
la. Throughout this procedure, unless otherwise
stated, the term water means distilled water.
la. Consult the manufacturer's manual for assistance
in correcting any malfunction.
V.A.I.1




Page No. 15-9
-------
EFFLUENT MONITORING PROCEDURE: Titrimetric Determination of Total Residual Chlorine in Wastewater Effluents
—-—————————	Page No. 15-10
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
B. Reagent Preparation





1. Starch Indicator
1.
2.
Weigh 5 g of soluble
starch.
Transfer it to a mortar.
la.
Use a trip balance.


3.
Measure 1 liter of water.
3a.
Use a 1 liter graduated cylinder.


4.
Pour the water into a
2 liter Erlenmeyer flask.




5.
Bring the water to a boil.
5a.
5b.
Use a hot plate.
While the water comes to a boil, do steps 6 & 7.


6.
Add 1 ml of water to the
starch in the mortar.
6a.
Use a 10 ml graduated cylinder to measure the
water.


7.
Grind the starch and water
together.
7a.
7b.
7c.
Use a pestle.
The objective is to form a thin paste.
A few additional drops of water may have to be
added.


8.
Slowly pour the thin paste
into the boiling water.
8a.
8b.
Be cautious about the hot flask.
If the water has not yet come to a boil, wait
until it does.


9.
Invert a 250 ml beaker and
place it on top of the
Erlenmeyer flask.
9a.
As protection against contamination.

1
10.
Turn the hot plate off.



-------
EFFLUENT MONITORING PROCEDURE: Titrimetric Determination of Total Residual Chlorine in Wastewater Effluents
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
B. Reagent Preparation
(continued)
2. Potassium Iodide,
KI, 10%
11.	Remove the flask from
the hot plate.
12.	Allow the starch to
stand overnight.
13.	Carefully decant the
supernatant liquid into a
1 liter glass-stoppered
bottle.
14.	Weigh 1.25 g of salicylic
acid.
15.	Add it to the bottle.
16.	Swirl the bottle.
1.	Weigh 10 g of potassium
iodide, KI
2.	Transfer it to a 250 ml
Erlenmeyer flask.
3.	Measure 90 ml of water.
4.	Add it to the flask.
11a. Caution: the flask is hot.
12a. Prepare the other reagents while the starch
solution is standing.
14a. Use an analytical balance (or trip balance if it
weighs to the second decimal place).
16a. To dissolve the salicylic acid.
la. Fourteenth Standard Methods does not specify the
strength of this solution. Ten % is the author's
opinion.
3a. Use a 100 ml graduated cylinder.
Page No. 15-11
-------
EFFLUENT MONITORING PROCEDURE: Titrimetric Determination of Total Residual Chlorine in Wastewater Effluents
"	Page No. 15-12
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
B. Reagent Preparation
(continued)
3. Standard Arsenite,
0.1N
5.	Swirl the flask.
6.	Transfer the solution to
a 100 ml glass-stoppered
bottle.
1.	Wipe a weighing bottle
with a tissue.
2.	Weigh about 5 g of arsenic
trioxide, As^O., in the
weighing bottle.
3.	Remove the bottle from
the balance.
4.	Fill a 1 liter volumetric
flask about one-half full
of water.
5.	Place a small powder funnel
into the mouth of the flask
6.	Remove the top of the
weighing bottle.
7.	Carefully turn the bottle
upside down into the funnel
in the volumetric flask.
5a. To dissolve the potassium iodide.
la. To remove fingerprints.
lb. Throughout B.3., always handle the bottle with
a tissue so as to avoid fingerprints.
2a. To four places to the right of the decimal.
2b. Arsenic trioxide, AspO;}, is extremely toxic.
Wipe up any spilled powder with a damp tissue,
discard the tissue, and thoroughly wash your
hands with soap and water.
7a. The arsenic trioxide is powdery, and will tend
to "fly around"; so do this step carefully.

-------
EFFLUENT MONITORING PROCEDURE: Titrimetric Determination of Total Residual Chlorine in Wastewater Effluents
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
B. Reagent Preparation
(continued)
8.
Gently tap the bottom and
sides of the weighing
bottle.
8a. So as to knock more of the arsenic trioxide into
the funnel.
8b. Some of the solid will stay in the bottle.


9.
Remove the weighing bottle
from the funnel.
9a. Remember the arsenic trioxide toxicity when you
later wash the bottle.


10.
Replace the top of the
weighing bottle.



11.
Reweigh the weighing
bottle.
11a. Use the same analytical balance that you used
before.
lib. Record the weight to four places to the right of
the decimal point.


12.
Using a plastic squeeze
bottle of water, carefully
wash the arsenic trioxide
dcwn into the volumetric
flask and remove the funnel
12a. Use a minimum of water.


13.
Measure 100 ml of water.
13a. Use a 100 ml graduated cylinder.


14.
Weigh 15 g of sodium
hydroxide, NaOH.
14a. Use a trip balance.


15.
Add the sodium hydroxide
and 100 ml of water to the
volumetric flask.



16.
Swirl the flask gently.
16a. To dissolve the sodium hydroxide and arsenic
trioxide.

Page No. 15-13
-------
,EF^LJJ^tT_WDNJ[TiOR>I^G__PiRO^EDyR^: Titrimetric Determination of Total Residual Chlorine in Wastewater Effluents
Page No. 15-14
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
Reagent Preparation
(continued)
17.	Add 150 ml of water to
the flask.
18.	Gently swirl the flask.
19.	Insert the glass tube
leading from the lecture
bottle of carbon dioxide,
C02, into the volumetric
flask; see the figure
on page
20.	Carefully open the lecture
bottle valve and adjust
the flow of carbon dioxide
so that about 1 bubble
per second comes from the
end of the tube.
21.	Continue the addition of
carbon dioxide for about
15 minutes.
2 2. Remove the tube from the
flask.
2 3. Close the lecture bottle
valve.
2 4. Add water to the 1 liter
mark of the flask.
2 5. Thoroughly mix the con-
tents of the flask.
17a. Use a 100 ml graduated cylinder.
18a. To thoroughly mix the contents.
19a. The glass tube should extend to the bottom
of the flask.

-------
EFFLUENT MONITORING PROCEDURE: Titrimetric Determination of Total Residual Chlorine in Wastewater Effluents
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
B. Reagent Preparation
(continued)
4. Standard Iodine 0.1N
26.	Transfer the solution to
a 1 liter glass-stoppered
bottle.
27.	Calculate the strength
(normality, N) of the
arsenic trioxide.
1.	Weigh 40 g of potassium
iodide, KI.
2.	Transfer it to a 250 ml
Erlenmeyer flask.
3.	Measure 25 ml of water.
4.	Add the water to the flask
5.	Swirl the flask.
6.	Weigh 13 g of resublimed
iodine, I2-
26a. The arsenic trioxide solution is also extremely
toxic. If any is spilled on the skin, rinse it
off immediately with large amounts of tap water.
27a. N = A - B
49.455
N = the strength (normality, N) of the arsenic
trioxide.
A = the weight, in g, of the weighing bottle
+ top + arsenic trioxide.
B = the weight, in g, of the weighing bottle
+ top + arsenic trioxide residue.
27b. The arsenic trioxide solution is stable almost
indefinitely.
la. Use a trip balance.
3a. Use a 100 ml graduated cylinder.
5a. To dissolve the potassium iodide.
6a. Use a trip balance.
Page No. 15-15
-------
EFFLUENT MONITORING PROCEDURE: Titrimetric Determination of Total Residual Chlorine in Wastewater Effluents
Page No. 15-16
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
Reagent Preparation
(continued)
7.	Add it to the Erlenmeyer
flask.
8.	Swirl the flask.
9.	Transfer the solution to
a 1 liter volumetric
flask.
10.	Fill the flask to the 1
liter mark with water.
11.	Thoroughly mix the con-
tents of the flask.
12.	Transfer the solution to
a 1 liter glass-stoppered
bottle.
8a. To dissolve the iodine.
9a. Rinse the Erlenmeyer flask with several small
portions of water and add the rinsings to the
volumetric flask.
12a. The strength of this solution is approximately
0.1N.

-------
EFFLUENT MONITORING PROCEDURE: Titrimetric Determination of Total Residual Chlorine in Wastewater Effluents
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
Reagent Preparation
(continued)
5.	Sodium Hydroxide,
NaOH, 0.3N
6.	Hydrochloric Acid,
HC1, 6 N
1.	Weigh 12 g of sodium
hydroxide, NaOH.
2.	Transfer it to a 2 liter
Erlenmeyer flask.
3.	Measure 1 liter of water.
4.	Add the water to the flask.
5.	Swirl the flask.
6.	Transfer the 0.3N base to
a 1 liter plastic bottle.
1.	Measure 50 ml of water.
2.	Pour it into a 250 ml
Erlenmeyer flask.
3.	Measure 50 ml of 12 N
hydrochloric acid, HC1
4.	Pour it slowly into the
flask.
5.	Thoroughly mix the contents
of the flask.
la. Use a trip balance.
3a. Use a 1 liter graduated cylinder.
5a. To dissolve the sodium hydroxide.
la. Use a 100 ml graduated cylinder.
3a. In a well ventilated area.
3b. Use a 100 ml graduated cylinder.
The usual concentration of hydrochloric acid as
it is purchased for ordinary laboratory use is
12N. More dilute concentrations can be pur-
chased, however. Twelve N acid can be detected
by gently blowing across the open bottle top, the
formation of white fumes indicates that the hydro-
chloric acid is 12 N. The more dilute concentra-
tions do not fume.

Page No. 15-17
-------
EFFLUENT MONITORING PROCEDURE: Titrimetric Determination of Total Residual Chlorine in Wastewater Effluents
Page No. 15-18
OPERATING PROCEDURES
STEP SEQUENCE
INFORMAT ION/0 P E RAT ING GOAL S/SPECIFICATIONS
TRAINING
GUIDE NOTES
Reagent Preparation
(continued)
7. Standard Phenyl-
arsine oxide (PAO),
0.01N
6. Transfer the 6N acid to a
glass-stoppered bottle.
1. Weigh 0.8 g of phenylarsine
oxide (PAO) powder.
2.	Transfer it to a 250 ml
Erlenmeyer flask.
3.	Measure 150 ml of 0.3N
sodium hydroxide.
4.	Pour it into the flask.
5.	Stir the contents of the
flask.
6.	Turn off the magnetic
stirrer.
7.	Allow any solid material
remaining in the flask to
settle.
8.	Decant 110 ml of the
supernatant liquid into a
250 ml graduated cylinder.
9.	Measure 950 ml of water.
10. Pour it into a 2 liter
Erlenmeyer flask.
la. Use a trip balance.
lb. Phenlarsine oxide (PAO) is extremely toxic.
Wipe up any spilled powder with a damp tissue,
discard the tissue, and thoroughly wash your
hands with soap and water.
3a. Use a 100 ml graduated cylinder.
5a. By means of a magnetic stirrer.
5b. Until the PAO dissolves. It may take a long time,
7a. For 30 minutes.
9a. Use a 1 liter graduated cylinder.
-------
EFFLUENT MONITORING PROCEDURE: Titrimetric Determination of Total Residual Chlorine in Wastewater Effluents
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
B. Reagent Preparation
(continued)
1.	Place a mark at the 950 ml
level.
2.	Pour 150 ml of the water
back into the graduated
cylinder.
3.	Discard the 150 ml of
water.
4.	Pour the 110 ml of super-
natant PAO into the 800
ml of water in the 2 liter
Erlenmeyer flask.
5.	Swirl the flask.
6.	Standardize a pH meter
with a pH 7 buffer.
7. Measure the pH of the
solution.
18. Add a few drops of the
6N hydrochloric acid to
the flask.
9. Swirl the flask.
20.	Measure the pH of the
solution.
21.	Repeat steps 18 through 20
above until the pH of the
solution is between 6.0
and 7.0.
15a. To thoroughly mix the contents.
17a. It should be between 6.0 and 7.0. If it is not,
do steps 18 through 20. If it is, proceed to
step 22.
18a. Use an eyedropper.
19a. To thoroughly mix the contents.
Page No. 15-19
-------
EFFLUENT MONITORING PROCEDURE: Titrimetric Determination of Total Residual Chlorine in Wastewater Effluents
Page No. 15-20
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
B. Reagent Preparation
(continued)
8. Acetate Buffer
Solution, pH 4.0
22.	Add enough water to the
flask to bring the level
to the 950 ml mark.
23.	Thoroughly mix the contents
of the flask.
24.	Transfer the solution to
a 1 liter glass-stoppered
bottle.
1.	Measure 400 ml of water.
2.	Pour it into a 1 liter
volumetric flask.
3.	Weigh 146 g of anhydrous
sodium acetate, NaC2H302.
4.	Transfer it to the flask.
5.	Swirl the flask.
6.	Measure 457 ml of acetic
acid, HC2H302.
7.	Add it to the flask.
8.	Add water to the 1 liter
mark.
9.	Swirl the flask.
10. Transfer the solution to
a 1 liter glass-stoppered
bottle.
22a. The PA0 solution is also extremely toxic. If
any is spilled on the skin, rinse it off im-
mediately with large amounts of tap water.
la. Use a 500 ml graduated cylinder.
3a. Use a trip balance.
3b. Two hundred forty-three g of sodium acetate
trihydrate, NaC2H.j02-3H20 may also be used.
5a. To dissolve the solid.
6a. Use a 500 ml graduated cylinder.
6b. In a well ventilated area.
9a. To thoroughly mix the contents.
-------
EFFLUENT MONITORING PROCEDURE: Titrimetric Determination of Total Residual Chlorine in Wastewater Effluents
OPERATING PROCEDURES
C. Standardization of
Reagents
1. Standardization of
the iodine.
STEP SEQUENCE
1.	Pipet 50.0 ml of the
standard arsenite (B.3.)
into a 250 ml Erlenmeyer
Flask.
2.	Measure 2 ml of starch;
add it to the flask and
mix.
3.	Fill a 50 ml buret with
the approximately 0.1N
iodine (B.4.).
4.	Remove air bubbles from
the buret tip.
5.	While swirling the flask
(or using a magnetic
stirrer), add iodine from
the buret to the flask at
a fast dropwise rate.
5. Whem you see a blue color
forming where the drops
of iodine hit the liquid
in the flask, stop the
addition of the iodine.
7.	Add 3 drops of concentra-
ted hydrochloric acid to
the flask.
8.	Swirl the flask.
INFORMATION/OPERATING GOALS/SPECIFICATIONS
la. Use a 50.0 ml volumetric pipet.
2a. Use a 10 ml graduated cylinder.
3a. Use a 10 ml graduated cylinder.
5a. Constant and thorough mixing is important.
6a. Even though a localized blue color formed, the
overall solution should still be colorless when
the solution is mixed.
6b. If it is blue, you have added too much iodine.
Rinse out the flask and start again at step
C.l.l. above.
7a. Use an eyedropper.
8a. To thoroughly mix the contents.
8b. Bubbles of carbon dioxide will form.
TRAINING
GUIDE NOTES
Page No. 15-21
-------
iEF^^^^£^__MOn^TO|R^NG_
-------
EFFLUENT MONITORING PROCEDURE: Titrimetric Determination of Total Residual Chlorine in Wastewater Effluents
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
C. Standardization of
Reagents (continued)
2.	Measure 100 ml of water.
3.	Pour it into a 1 liter
volumetric flask.
4.	Weigh 25 g of potassium
iodide, KI.
5.	Add the potassium iodide,
KI, to the flask.
6.	Swirl the flask.
7.	Measure the calculated
volume of iodine to be
diluted to 1 liter to ob-
tain a 0.0282 N solution.
8.	Add it to the volimetric
flask containing the
potassium iodide solution.
9.	Add water to the 1 liter
mark.
10.	Mix the contents of the
flask thoroughly.
11.	Transfer the 0.0282 N
iodine solution to a 1
liter glass-stoppered
bottle which has been
wrapped in aluminum foil.
2a. Use a 100 ml graduated cylinder.
4a. Use a trip balance.
6a. To dissolve the potassium iodide, KI.
7a. E will be an "unusual" volume, about 282 ml. Use
a 500 ml graduated cylinder to measure to the
nearest 10.0 ml, and a 10 ml graduated pi pet to
measure to the nearest 1.0 ml. For example,
measure 280 ml in the cylinder, and 2.0 ml in
the pipet.
11a. The solution should be protected from the sun-
light.
lib. This solution should be standardized on each day
it is used.
lie. The standardization is carried out exactly as
described in section C.l. above except: pi pet
10.0 ml of arsenic trioxide instead of 50.0 ml,
and use a 125 ml Erlenmeyer flask instead of a
250 ml flask. Also, the diluted iodine (0.0282 N)
is use(J ii) the buret, instead of the approximately
°-1 N 10d1m-	Page No. 15-23
-------
EFFLUENT MONITORING PROCEDURE: Titrimetric Determination of Total Residual Chlorine in Wastewater Effluents
Page No. 15-24
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
C. Standardization of
Reagents (continued)
3. Standardization of
the PAO
1.	Pipet 10.0 ml of freshly
standardized diluted
iodine titrant (C.2.11.lie.)
into a 125 ml Erlenmeyer
flask.
2.	Fill a buret (B.7) with
the PAO.
3.	Remove air bubbles from
the buret tip.
4.	Measure 1.0 ml of potassium
iodide.
5.	Add it to the flask.
6.	Swirl the flask.
la. Use a 10 ml volumetric pipet.
4a. Use a 10 ml graduated cylinder.
6a. To thoroughly mix the contents.
6b. The solution is red brown in color.

-------
EFFLUENT MONITORING PROCEDURE: Titrimetric Determination of Total Residual Chlorine in Wastewater Effluents
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
Standardization of
Reagents (continued)
7.	While swirling the flask
(or using a magnetic
stirrer), add PAO from the
buret to the flask at a
fast dropwise rate.
8.	When the color of the
solution changes to a pale
yellow color, stop the
addition of PAO.
9.	Measure 2.0 ml of starch.
10.	Add the starch to the
flask.
11.	Swirl the flask.
12.	While swirling the flask,
(or using a magnetic
stirrer), continue addi-
tion of the PAO at a slower
dropwise rate than before.
3. Calculate the strength
(normality, N) of the PAO
7a. Constant and thorough mixing is important.
9a. Use a 10 ml graduated cylinder.
11a. To thoroughly mix the contents.
lib. The solution is now medium to pale blue in color.
12a. About one-half as fast.
12b. At some point in the titration 1 drop of PAO will
cause the solution to turn colorless.
12c. Immediately stop the addition.
12d. Record the ml of PAO used.
13a. N = H x 10.0
I
N = the strength (normality, N) of the PAO; it
will be approximately 0.01.
H = the normality of the iodine (C.2.11.lie.)
I = the ml of PAO used from the buret

Page No. 15-25
-------
MONITORING PROCEDURE: Titrimetric Determination of Total Residual Chlorine in Wastewater Effluents
Page No. 15-26
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
C. Standardization of
Reagents (continued)
4. PAO Solution
0.00564N
Checking the N of
the diluted PAO
1. Calculate the volume of
PAO (C.3.13.13a.) to be
diluted to 1 liter to ob-
tain a 0.00564 N solution.
2. Measure the calculated
volume of PAO to be
diluted to 1 liter to ob-
tain a 0.00564N solution.
3.	Add the measured volume to
a 1 liter volumetric flask.
4.	Add water to the 1 liter
mark.
5.	Thoroughly mix the con-
tents of the flask.
1. Repeat steps C.3.1.
through C.3.13.
la. J = K x 1000
L
J = ml of PAO (C.3.13.13a.) to be diluted
K = desired N of the PAO solution after
dilution, 0.00564.
1000 = desired volume (in ml) of the diluted PAO
L = N of PAO (C.3.13.13a.)
2a. J will be an "unusual" volume; about 500 ml. If
J is more than 500 ml, use a 1 liter graduated
cylinder to measure to the nearest 10.0 ml. If
0 is less than 500 ml, use a 500 ml graduated
cylinder to measure to the nearest 10.0 ml.
After using the appropriate cylinder, use a 10 ml
graduated pipet to measure to the nearest 1.0 or
0.1 ml.
5a. If the dilution was done properly, the N of the
PAO is 0.00564 N.
la. Except that the diluted PAO is added from the
buret, instead of the stronger PAO.
lb. The N of the PAO is 0.00564.
1c. If it is not, discard the diluted PAO and repeat
sections C.£, C.3., and C.4.
-------
EFFLUENT MONITORING PRQCEDURF: Titrimetric Determination of Total Residual Chlorine in Wastewater Effluents
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
Procedure
1.
Pi pet 5.0 ml of the
0.00564 N PAO (C.5.) into
a 30 ml beaker.
la. Use a 5 ml volumetric pipet.


2.
Weigh 1.0 g of potassium
iodide, KI.
2a.
Use a trip balance.


3.
Add 1t to the beaker.




4.
Stir the beaker contents.
4a.
4b.
Use a stirring rod.
To dissolve the solid.


5.
Standardize a pH meter with
a pH 4 buffer.




6.
Check the pH of the solu-
tion in the beaker.
6a.
6b.
It must be between 3.5 and 4.2 before the
titration is begun.
If it is, proceed to step 11. If it is not, do
steps 7 through 10.


7.
Add 5-10 drops of acetate
buffer.
7a.
Use an eyedropper.


8.
Swirl the beaker.
8a.
To thoroughly mix the contents.


9.
Recheck the pH.
9a.
It must be between 3.5 and 4.2 before the
titration is begun.


10.
Repeat steps 7, 8, and 9
above until the pH is be-
tween 3.5 and 4.2.




11.
Pour the contents of the
beaker into a 500 ml
Erlenmeyer flask.



Page No. 15-27
-------
EFFLUENT MONITORING PROCEDURE: Titrimetric Determination of Total Residual Chlorine in Wastewater Effluents
Page No. 15-28
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
Procedure
(continued)
12. Measure 200 ml of sample.
13.	Rinse the 30 ml beaker
with several portions of
sample from the graduated
cylinder.
14.	Pour the rest of the
sample into the flask.
15.	Swirl the flask.
16.	Fill a 5 ml buret with
the 0.0282 N iodine
(C.2.11.11c.)
17.	Remove air bubbles from
the buret tip.
18.	Measure 1.0 ml of starch.
19.	Add it to the flask.
20.	Swirl the flask.
21.	While swirling the flask
(or using a magnetic
stirrer), add the iodine
from the buret to the flask
at a dropwise rate.
12a. There can be no delay between the time the sample
is collected and the time the analysis is done.
Protect the sample from sunlight and do not
agitate it.
12b. Use a 250 ml graduated cylinder.
12c. Two hundred ml of sample are used when the ex-
pected concentration of total residual chlorine
is less than 10 mg/1.
12d. One hundred fifty ml would be used if the ex-
pected concentration is between 10 and 15 mg/1.
12e. One hundred ml would be used if the expected
concentration is between 15 and 20 mg/1.
13a. Pour each rinse into the Erlenmeyer flask.
15a. To thoroughly mix the contents.
18a. Use a 10 ml graduated cylinder.
20a. To thoroughly mix the contents.
21a. About one drop per second.
21b. Thorough and cQnstant mixing are essential dur-
ing tne titration.
-------
EFFLUENT MONITORING PROCEDURE: Titrimetric Determination of Total Residual Chlorine in Wastewater Effluents
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
D.	Procedure
(continued)
E.	Calculation
22.	At some point the addi-
tion of one drop of iodine
will cause formation of a
blue color.
23.	Immediately stop the
addition of iodine when
this happens.
24.	Record the ml of iodine
used from the buret.
1. Calculate the total
residual chlorine con-
tent of the sample in
mg/1.
22ek The color will not fade on standing for a few
seconds.
24a. To the nearest 0.1 ml.
la. mg of total residual chlorine per liter of
sample = (5.0 - 5 A) X 200
B
lb. A = ml of iodine used from the buret.
B = ml of sample
1c. When B = 200, mg of total residual chlorine per
liter of sample = (5.0 - 5 A).

Page No. 15-29
-------
EFFLUENT MONITORING PROCEDURE: Titrimetric Determination of Total
Residual Chlorine in Wastewater
TRAINING GUIDE
SECTION	TOPIC
I	Introduction
II	Educational Concepts - Mathematics
III	Educational Concepts - Science
IV	Educational Concepts - Communications
V*	Field & Laboratory Equipment
VI	Field & Laboratory Reagents
VII	Field & Laboratory Analysis
VIII	Safety
IX	Records & Reports
~Training guide materials are presented here under the headings marked *.
Page No. 15-30
-------
EFFLUENT MONITORING PROCEDURE: Tltrimetric Determination of Total Residual
Chlorine in Wastewater
FIELD & LABORATORY EQUIPMENT	Section V

TRAINING GUIDE NOTE
REFERENCES/RESOURCES
A. 1.1
If the glassware is especially dirty and cannot be
cleaned with ordinary detergents, chromic acid
cleaning may be required.
Standard Methods, 14th ed
1975, p. 336, par. 2.C.2

1.
Pour 35 ml of distilled water in a 250 ml beaker,


2.
Add about 1/8 teaspoon (simply estimate this
quantity) of sodium dichromate, Ma^Cr^O^
to the water.


3.
Swirl the beaker until the sodium dichromate
has dissolved.


4.
Keep repeating steps 2 and 3 until no more
sodium dichromate will dissolve.


5.
Pour the solution into a 2 liter beaker.


6.
Slowly pour 1 liter of concentrated sulfuric
acid, HgSO^, Into the 2 liter beaker.
CAUTION: Use eyeglasses and protective clothing.


7.
Stir the mixture thoroughly.


8.
Store it in a glass stoppered bottle.


9.
The cleaning solution should be at a temperature
of about 50°C when 1t 1s used.


10.
It may therefore be necessary to warm the
cleaning solution.


11.
When using the warm cleaning solution, fill the
piece of glassware with the solution.


12.
Allow it to soak for 2-3 minutes (or longer).


13.
Pour the cleaning solution back Into the storage
bottle.


14.
Rinse the piece of glassware ten times with
tap water.


15.
The cleaning solution may be reused until 1t
turns green.


16.
It should then be discarded.

Page No. 15-31
-------
A PROTOTYPE FOR DEVELOPMENT OF
ROUTINE OPERATIONAL PROCEDURES
for the
DETERMINATION OF TOTAL SUSPENDED
(NON-FILTERABLE) SOLIDS, mg/liter
as applied in
WASTEWATER TREATMENT FACILITIES
and in the
MONITORING OF EFFLUENT WASTEWATERS
Developed by the
National Training and Operational Technology Center
Municipal Operations and Training Division
Office of Water Program Operations
U.S. Environmental Protection Agency
CH.SOL.SUS.EMP.la.6.77
Page No. 16-1
-------
EFFLUENT MONITORING PROCEDURE: Total Suspended (Non-Filterable) Solids, mg/liter
This Operational Procedure was developed by:
Name	Audrey D. Kroner
Address EPA, OilPO, NTOTC, Cincinnati, Ohio *5263
Position Chemist-Instructor
Education and Technical Background
B.A. Edgecliff College
1 year Industrial Research Chemist
8 years Secondary School Chemistry Instructor
4 years DHEW-DI Water Quality Program Chemist
7-1/2 years DI-EPA Chemist-Instructor
Page No. 16-3
-------
EFFLUENT MONITORING PROCEDURE: Total Suspended (Non-Filterable) Solids,
mg/liter
1.	Objective:
To determine total suspended (non-filterable) solids on a weight (mg/liter)
basis.
2.	Brief Description of Analysis:
A well-mixed sample is filtered through a weighed, standard glass fiber filter
disc in a filtration assembly. The filter disc with retained residue is dried
in an oven at 103 - 105 C until a constant weight is obtained. The difference
between the weight of the filter disc plus residue (g) and the original weight
of the filter disc (g) is divided by the milliliters of sample filtered, then
multiplied by 1,000,000. The final result is recorded as total suspended (non-
filterable) solids, mg/liter.
3.	Applicability of this Procedure:
a.	Range of Concentration:
10 to 20,000 mg/liter
b.	Pretreatment of Sample:
The Federal Register Guidelines do not specify any pretreatment.
c.	Treatment of Interferences in Samples:
This procedure includes directions and information about choosing a sample
volume small enough to prevent getting too much residue on the filter.
(This entraps water and prolongs drying periods.)
*
No other interferences are noted in the Source of Procedure .
~Source of Procedure: Methods for Chemical Analysis of Water and Wastes, 1974,
U.S. Environmental Protection Agency, Methods Development & Quality Assurance
Research Laboratory, Cincinnati, Ohio, 45268( p. 268.
Page No. 16-4
-------
EFFLUENT MONITORING PROCEDURE: Total Suspended (Non-Filterable) Solids,
mg/liter
Operating Procedures:
A. Prepare the filter disc
60 minutes in oven at 103°-105°C
20-30 minutes in a desiccator
B.	Prepare to test the sample
C.	Weigh the filter disc
D.	Seat the filter disc
E.	Filter the sample
F.	Wash down walls of filter apparatus
G.	Dry filter disc and residue
J.I. Clean the filtration equipment
60 minutes in oven at 103°-105°C
	20-30 minutes in a desiccator
H.	Weigh filter disc and residue
I.	Check for complete drying
30 minutes in oven at 103°-105°C
	20-30 minutes in a desiccator
Finish check for complete drying
0.2 Clean filter disc support
K. Calculate total suspended (non-filterable) solids, mg/liter
L. Report the data
Page No. 16-5
-------
EFFLUENT MONITORING PROCEDURE: Total Suspended (Non-Filterable) Solids,
mg/liter
Equipment and Supply Requirements
A. Capital Equipment:
Balance, analytical, capable of weighing to 0.1 mg under a 200 g load
Oven, drying, for use at 103°-105°C.
Vacuum source or pump drawing 15 inches mercury
B. Reusable Supplies:
1 Cylinder, graduated, 25 or 50 ml
1 Cylinder, graduated, volume equal to or greater than the volume of
sample to be filtered. (100 ml is commonly used. For sample
volumes less than 10 ml, a wide-tip pipet can be used with a
pi pet bulb to draw sample into pipet.)
1 Oesiccator (for storing filter discs on watch glasses, etc.)
1 Flask, suction, with side arm, 1000 ml
1 Hose connection from suction flask to vacuum source
1 Pinchcock clamp to use on hose
1 Filter holder: membrane filter holder assembly or Buchner funnel or
Hlrsch funnel. The filter holder should have a stopper which fits
into the mouth of the 1000 ml suction flask. Gooch crucibles may
be used—one for each sample plus one adapter to hold the crucibles
1n the mouth of the 1000 ml suction flask.
1 Support for filter disc during drying (watch glasses, etc., number
depends on number of samples). If Gooch crucibles are used, omit
this Item.
1 Pair Tongs or gloves, etc., to remove crucibles or watch glass from
the oven
1 Pair Forceps (flat, to handle filter discs)
1 Wash Bottle, squeeze type for distilled water
1 Set Cork Borers
Page No. 16-6
-------
EFFLUENT MONITORING PROCEDURE: Total Suspended (Non-Filterable) Solids,
mg/liter
C. Consumable Supplies:
Filter discs, glass fiber, without organic binder, Reeve Angel type
934A or 984H, Gel man type A, Whatman GF/C or equivalent. Diameter
should be large enough so disc will cover openings in the filter
holder to be used.
Marking ink to permanently mark glass or porcelain. A marking tool can
be used instead.
Notebook, bound
Tissues, soft (for balance work)
Water, distilled
Page No. 16-7
-------
EFFLUENT MONITORING PROCEDURE: Total Suspended (Non-Filterable) Solids, mg/liter
Page No. 16-8
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATION'S
TRAINING
GUIDE NOTES
TOTAL SUSPENDED (N0N-F1
A. Preparing the
Filter Disc
LTERABLE) SOLIDS, mg/liter
1.	Gather equipment.
2.	Place filter holder with
stopper or adapter into the
suction flask.
3.	Attach hose.
4.	Pick up a filter disc.
5.	Place filter disc on the
filter holder.
6.	Apply vacuum.
7.	Measure out about 20 ml
distilled water.
la. See page 6 for list of necessary equipment. The
oven should be turned on and set for 103°-105°C
temperature.
lb. Filter disc supports (watch glasses, etc.) or
Gooch crucibles should have permanent identifica-
tion marks.
lc. Be sure equipment is clean.
2a. Twist, pressing downward for air-tight fit.
3a. From side arm of suction flask to the vacuum
source.
4a. Using forceps.
5a. Wrinkled surface of filter disc facing upward.
5b. Disc should cover all openings in filter holder.
6a. Gradually, to seat the filter disc. A pinchcock
clamp on the vacuum hose can be used to regulate
application of vacuum.
6b. If a membrane filter holder is used, attach funnel
now and tighten the collar.
7a. In a 25 or 50 ml graduated cylinder.
I
(p. 25)
V.A.lb
(p. 271
V.A.lc
(p. 27)
-------
EFFLUENT MONITORING PROCEDURE: Total Suspended (Non-Filterable) Solids, mg/liter
DERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
Preparing the Filter
Disc (Continued)
8. Pour the 20 ml distilled water
on to the disc.
9. Measure out about 20 ml
distilled water.
10.	Pour this second 20 ml amount
of distilled water on to the
disc.
11.	Measure out about 20 ml
distilled water.
12.	Pour this third 20 ml amount
of distilled water on to the
disc.
13.	Continue vacuun application.
14. Turn off vacuum.
5. Loosen the filter disc from
the filter holder.
8a. Vacuum still being applied.
8b. To rinse off the filter disc.
8c. If fibers of disc form a lumpy area, discard the
disc and begin again at step 4.
9a. In the same graduated cylinder.
0a. Vacuum still being applied.
0b. A second rinse for the disc.
I la. In the same graduated cylinder.
12a. Vacuum still being applied.
12b. A third rinse for the disc.
13a. For 2 minutes to remove all traces of water.
13b. If a membrane filter holder is used, loosen the
collar and remove the funnel.
14a. Break vacuum by pushing upward on the rubber
adapter or stopper until air can enter the flask.
15a. If a Gooch crucible is used, omit this step.
15b. If a membrane filter holder is used, use forceps
to loosen the disc. Be careful not to damage the
disc.
Page No. 16-9
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EFFLUENT MONITORING PROCEDURE: Total Suspended (Non-Filterable) Solids, mg/liter
Paae No. 16-10
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
A. Preparing the Filter
Disc (Continued)
16.	Slide the filter disc onto
a suitable support.
17.	Put disc (on support) into
an oven.
18.	Remove disc (on support)
from oven.
19.	Allow disc (on support) to
cool partially to room
temperature.
20.	Put disc (on support) into
desiccator.
21.	Store disc in desiccator
until needed.
16a. If a Gooch crucible is used, remove the crucible
with the filter disc in it. Wipe the outside
with a tissue to remove droplets of water, finger-
prints, etc. Do not directly handle the crucible
during the procedure. Use tissue, forceps or
tongs instead.
16b. If a membrane filter holder is used, use a dry
watch glass, etc., to hold the disc.
16c. The filtration assembly can be left as is for
future use.
17a. To dry at 103°-105°C.
17b. For 30 minutes in a mechanical convection oven.
17c. For 60 minutes in a gravity convection oven.
17d. Note: Do not open oven door during drying period.
18a. With tongs or gloves, etc.
19a. Place on clean, heat-resistant surface for about
three minutes.
20a. With tongs or gloves, etc.
20b. Desiccant must be dry.
20c. Desiccator should be air-tight with enough room
so disc supports do not touch each other or the
side of the desiccator.
21a. Disc and support should be cooled to room tem-
perature before weighing—20 to 30 minutes.
V.A.20a.
(p. 27)




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EFFLUENT MONITORING PROCEDURE: Total Suspended (Non-Filterable) Solids, mg/liter
PERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
Preparing to Test
the Sample
1. Assemble filtering equipment
except for filter disc.
Record the sample identifica-
tion information.
la. Equipment list is on page 6.
lb. The filtering assembly used to prepare the disc
(rinsing it) can be re-used at this time.
1c. Rinse water can remain in suction flask.
Id. Filter holder or Gooch crucible adapter should be
tightly in mouth of suction flask,
le. The oven should be turned on and set for 103°-105°C
temperature.
2a. Sample should be at hand before continuing with
this test.
2b. Use a laboratory notebook.
2c. Record "identification", "type", "date and time
collected", and name of "sample collector."
VII.B.2a
(p. 23)
IX.B.2b
(P. 51)
IX.B.2c
(p. 51)
Weighing the Filter
Disc
1.	Bring forceps, record book
and pen to balance table.
2.	Zero the balance.
3.	Remove filter disc (on support)
from desiccator.
4. Record filter disc identifica-
tion.
la. Use an analytical balance.
3a. If a Gooch crucible is being used, use a tissue,
forceps or tongs to remove it from the desiccator.
It should contain a rinsed, dried filter disc.
4a. Gooch crucible number or watch glass number.
(Examples: C-12, WG-1)
4b. In laboratory notebook.
4c. In column of the sample for which this disc will
be used.
4d. Labeled "filter identification."
V.C.4a
(P.
27)
IX.C.4
(p. 32)
Page No. 16-11
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EFFLUENT MONITORING PROCEDURE: Total Suspended (Non-Filterable) Solids, mg/liter
Page No. 16-12
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
C. Weighing the Filter
Disc (Continued)
5.	Place filter disc on balance
pan.
6.	Weigh the filter disc.
7.	Record the weight.
8.	Remove the filter disc from
the oalance pan.
9.	Return all weights on the
balance to zero position.
5a. If a Gooch crucible is being used, use a tissue,
forceps or tongs to place it on the balance pan.
5b. If a membrane filter holder is being used, use
forceps tc slide the filter disc from the storage
support (watch glass, etc.) on to the pan.
6a. To four decimal places.
6b. For Gooch crucibles, you can save weighing time by
keeping a list of the numbered crucibles with their
approximate weights so you have a beginning weight
for this operation.
7a. In laboratory notebook,
7b. In column of the sample for which this disc will
be used.
7c. Labeled "weight of filter (g)." If Gooch crucibles
are used, this is the weight of the crucible con-
taining a filter disc.
8a. If a Gooch crucible is being used, use tissue,
forceps or tongs to remove crucible containing
filter disc.
8b. If a membrane filter holder is being used, use
forceps to slide the filter disc from the pan on
to its storage support (watch glass, etc.).
IX.C.7
(p. 32)




-------
EFFLUENT MONITORING PROCEDURE: Total Suspended (Non-Filterable) Solids, mg/liter
OPERATING PROCEDURES
). Seating the Filter
Disc
STEP SEQUENCE
1. Slide the filter disc on to
the filter holder held in the
mouth of the suction flask.
2. Apply vacuum.
3. Pour about 5 ml distilled
water on to the filter disc.
4. Leave vacuum on.
INFORMATION/OPERATING GOALS/SPECIFICATIONS
la. If a Gooch crucible is used, put crucible and disc
into the Gooch crucible adapter,
lb. If a membrane filter holder is used, place the
wrinkled surface of the disc facing upward on the
filter holder,
lc. If a series of funnel type filter assemblies are
used, be sure to write the filter disc identifica-
tion number on the corresponding funnel or flask.
2a. Gradually» to help seat filter disc. A pinchcock
clamp on the vacuum hose can be used to regulate
application of vacuum.
2b. If a membrane filter holder is being used, attach
funnel now and tighten the collar.
3a. Vacuum still being applied.
3b. Can use squeeze bottle of distilled water and
estimate volume.
3c. Wetting helps seat filter against holder.
TRAINING
GUIDE MOTES
E. Filtering the
Sample
1. Record date and time.
2. Select the volume of sample
to be filtered.
3. Shake the sample.
la. In laboratory notebook,
lb. In column of the sample to be filtered,
lc. Labeled "Date and Time Analysis Began."
2a. 100 ml of sample is a commonly used volume.
2b. CAUTION: Too much residue on the filter will entrap
water and may require prolonged drying. If sus-
pended solid concentration in the sample is
obviously great, choose a less-than-100 ml volume
of well mixed sample.
3a. So portion used is representative of all the sample,
IX.E.1
fo. 32)
VII.E.2b
(p. 29)
Page No. 16-13
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EFFLUENT MONITORING PROCEDURE: Total Suspended (Non-Filterable) Solids, mg/liter
Page No. 16-14
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
E. Filtering the
Sample (Continued)
4.	Immediately measure out the
selected volume.
5.	Pour the sample on to the
filter disc in the filtration
assembly.
6.	Rinse any sample left in the
cylinder on to the filter disc.
1. Leave suction on.
J. Record the total ml of sample
filtered.
4a. Using a graduated cylinder (use a wide tip pi pet
for volumes less than 10 ml).
4b. Measure rapidly since solids may settle in the
sample container while you are filling the cylinder.
4c. If you pour the sample to above the graduations,
pour that sample back into the bottle and begin
again at Step 3.
5a. You should filter all the sample you measure out
because you should rinse remaining, settled solids
out of the cylinder and on to the filter disc.
5b. If a series of samples are being filtered, be sure
you filter each sample through the filter disc you
weighed and designated for that sample on the lab
data sheet.
5a. With distilled water.
6b. As required.
6c. If suspended solid concentration on the filter disc
is obviously small, measure additional volumes of
well-mixed sample and filter these, rinsing the
cylinder each time.
3a. In laboratory notebook.
3b. In column of the sample filtered.
3c. Labeled "ml Sample Filtered."
IX.E.8
(P- 32)





-------
EFFLUENT MONITORING PROCEDURE: Total Suspended (Non-Filterable) Solids, mg/liter
PERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE .NOTES
Washing Walls of
Filter Apparatus
1. Rinse walls of filter holder
with about 10 ml distilled
water.
2.	Allow time for complete
drainage.
3.	Rinse walls of filter holder
with another 10 ml distilled
water.
4.	Allow time for complete
drainage.
5.	Rinse walls a third time with
about 10 ml distilled water.
6.	Continue vacuum application.
7. Turn off vacuum.
la. A squeeze bottle of distilled water can be used.
Estimate the 10 ml volume,
lb. Otherwise, use a graduate and direct the rinse onto
the walls,
lc. Suction should be applied.
3a. See information above for F.l.
5a. See information above for F.l.
5b. NOTE: The diluted filtrate cannot be used for a
dissolved solids determination.
6a. For two minutes to remove all traces of water.
6b. If a membrane filter holder is used, loosen the
collar and remove the funnel.
7a. Break vacuum by pushing upward on the rubber
adapter or stopper until air can enter the flask.
Page No. 16-15
-------
EFFLUENT MONITORING PROCEDURE: Total Suspended (Non-Filterable) Solids, mg/liter
Page No. 16-16
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
Drying the Filter
Disc and Residue
1. Loosen the filter disc from
the filter holder.
2. Slide the filter disc plus
residue on to its support.
3. Put disc (on support) into
an oven.
Remove disc (on support) from
oven.
Allow disc (on support) to
cool partially to room temper-
ature.
Put disc (on support) into
desiccator.
7. Allow time for disc to cool
to room temperature.
la. If a Gooch crucible is used, omit this step.
lb. If a membrane filter holder is used, use forceps
to loosen the filter disc.
2a. If a Gooch crucible is used, remove the crucible
with the filter disc in it. Wipe the outside with
a tissue to remove droplets of water, fingerprints,
etc. before drying.
2b. If a membrane filter holder is used, slide the
filter disc on to the same marked watch glass you
used earlier for its support.
3a. To dry at 103°-105°C.
3b. For 30 minutes in a mechanical convection oven.
3c. For 60 minutes in a gravity convection oven.
3d. NOTE: Do not open oven door during drying period.
3e. NOTE: While solids are in drying oven, do "J.
Cleaning the Equipment, Step 1."
4a. With tongs or gloves, etc.
4b. Let oven turned on and set for 103°-105°C tempera-
ture.
5a. Place on clean, heat-resistant surface for about
three minutes.
6a. Desiccant must be dry.
6b. Desiccator should be air-tight with enough room
so disc supports do not touch each other or the
side of the desiccator.
7a. Twenty to 30 minutes.
VII.G.3b
(P. 29)
V.G.6a
(p. 27)
-------
EFFLUENT MONITORING PROCEDURE: Total Suspended (Non-Filterable) Solids, mg/liter
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
H. Weighing the Filter
Disc and Residue
1.	Bring forceps, record book
and pen to balance table.
2.	Zero the balance.
3.	Remove filter disc dIus
residue (on support) from
desiccator.
4.	Place filter disc on balance
pan.
5. Weigh the filter disc plus
residue.
6. Record the weight.
la. Use the same analytical balance you used earlier
to weigh the disc.
3a. If a Gooch crucible is being used, use a tissue,
forceps or tongs to remove it from the desiccator.
4a. If a Gooch crucible is being used, use a tissue,
forceps or tongs to place it on the balance pan.
4b. If a membrane filter holder is being used, use
forceps to slide the filter disc from the storage
support (watch glass, etc.) onto the pan.
5a. To four decimal places.
5b. Use the "weight of the filter" (or of the Gooch
crucible with filter) on your Laboratory Data
Sheet as a beginning weight.
7. Remove the filter disc from
the balance pan.
8. Return all weights on the
balance to zero position.
6a.
6b.
6c.
7a.
7b.
In laboratory notebook,
In column of the sample for which the disc was used.
Labeled "1st weight of filter plus residue (g)."
If Gooch crucibles are used, this is the weight of
the crucible containing a filter disc with residue.
If a Gooch crucible is being used, remove crucible
containing filter disc with residue.
If a membrane filter holder is being used, use
forceps to slide the filter disc with residue back
on to its support (watch glass, etc.)
IX.H.6
(P- 32)
Page No. 16-17
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EFFLUENT MONITORING PROCEDURE: Total Suspended (Non-Filterable) Solids, mg/liter
Page No. 16-18
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
I. Check for Complete
Dryi ng
Put disc plus residue (on
support) into an oven.
2. Remove disc (on support)
from oven.
3.	Allow disc (on support) to
cool partially to room
temperature.
4.	Put disc (on support) into
desiccator.
5.	Allow time for disc to cool
to room temperature.
6.	Bring forceps, record book
and pen to balance table.
7.	Zero the balance.
8.	Remove filter disc (on
support) from desiccator.
9.	Place filter disc on
balance pan.
10. Weigh the filter disc plus
residue.
la. At 103°-105°C
lb. For 30 minutes.
lc. NOTE: Do not open oven door during drying period.
9a.
9b,
VII.I
(p. 30)
2a. Using tongs or gloves, etc.
Let oven i
perature.
2b. Let oven turned on and set for 103°-105°C tem-
3a. Place on clean, heat-resistant surface for about
three minutes.
4a. Desiccant must be dry.
4b. Desiccator should be air-tight with enough room
so disc supports do not touch each other or the
side of the desiccator.
5a. Twenty to 30 minutes.
6a. Use an analytical balance.
8a. If a Gooch crucible is being used, use a tissue,
forceps or tongs to remove it from the desiccator.
If a Gooch crucible is being used, use a tissue,
forceps or tongs to place it on the balance pan.
If a membrane filter holder is being used, use
forceps to slide the filter disc from the storage
support (watch glass, etc.) on to the pan.
10a. To four decimal places.
10b. Use the "1st weight of the filter plus residue
(g)" recorded on your data sheet for this sample
as a beginning weight.
-------
EFFLUENT MONITORING PROCEDURE: Total Suspended (Non-Filterable) Solids, mg/liter
rRAinii;
GUIDE NOTES
INFORMATION/OPERATING GOALS/SPECIFICATIONS
STEP SEQUENCE
OPERATING PROCEDURES
I. Check for Complete
Drying (Continued)
11. Record the weight.
12. Remove the filter disc plus
residue from the balance pan.
13.	Return all weights on the
balance to zero position.
14.	Find the difference between
the 1st and 2nd weights of
the filter plus residue.
15. Inspect the difference for
acceptable agreement of
these two weights.
11a. In laboratory notebook.
lib. In column of the sample for which the disc was
used.
lie. Labeled "2nd weight of filter plus residue (g)."
If Gooch crucibles are used, this is the weight
of the crucible containing a filter disc with
residue.
12a. If a Gooch crucible is being used, remove crucible
containing disc with residue. Save this.
12b. If a membrane filter holder is being used, use
forceps to slide the filter disc with residue back
on to its support (watch glass, etc.). Save this.
14a. In laboratory notebook.
14b. In column of the sample for which the disc was
used.
14c. Labeled "difference (lst-2nd)."
15a. if the weights agree, drying was complete so the
procedure is finished.
1)	The weights should ideally be constant £the
same weight, ± the possible balance error of
O.OOOlg (0.1 mg^|» Use the last weight obtained.
2)	An acceptable difference between these succes-
sive weights is no more than 0.0005g (0.5 mg).
In this case, use the last weight obtained for
the "final weight of filter plus residue (g)"
on line 13 of the Laboratory Data Sheet.
(Continued)
IX.1.11
(P. 32)
IX.I.14
(P. 32)
II.1.15a. 1
(p. 26)
II .1,15a.2
(p. 26)
Page No. 16-19
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EFFLUENT MONITORING PROCEDURE: Total Suspended (Non-Filterable) Solids, mg/liter
Page No. 16-2D
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
I. Check for Complete
Drying (Continued)

l
15b. If the weights do not meet the requirements of
agreement, repeat this "Section I: Check for Com-
plete Drying" until you do obtain two successive
weights that agree according to a.1) or a.2)
above. Use the last weight obtained as the "final
weight of filter plus residue (g)" on line 13 of
the Laboratory Data Sheet.
IX.1.15
(p. 32)

16.	Sign the laboratory data
sheet.
17.	Turn oven off.
16a. in laboratory notebook.
16b. In column for sample(s) you tested.
16c. Labeled "analyst."
IX.1.16
(p. 32)

18. Discard the filter disc plus
residue.
18a. Unless there is some reason for saving the solids.
18b. The filter disc support should be cleaned
according to "J. Cleaning the Equipment, Step 2."

J. Cleaning the Equip-
ment
1. Clean the filtration equip-
ment as soon as possible
after use (See G.3e).
la. Membrane filter holder assembly: Leave disc support
in suction flask, use squeeze bottle of distilled
water, rinse disc support while applying gentle
suction. Assembly need not be completely dry before
re-use.
lb. Hirsch funnel or Buchner funnel: Leave funnel in
suction flask and rinse with distilled water as
described above in J.1.1a.
1c. Gooch adapter: Leave in suction flask and rinse
the small glass funnel with distilled water
(squeeze bottle) while applying gentle suction.
Adapter need not be completely dry before re-use.
Id. Suction flask: Remove the rinsed filter holder.
Empty the flask through the top (not the side-arm).
Rinse it with tap water. Flask need not be com-
pletely dry before re-use.
(Continued)

-------
EFFLUENT MONITORING PROCEDURE: Total Suspended (Non-Filterable) Solids, mg/liter
PERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
J. Cleaning the Equip-
ment (Continued)
Clean the filter disc support
as soon as possible after use.
(See I.17.b)
le. Graduated cylinders: Rinse with distilled water.
These should be dry before re-use.
If. If stronger cleaning measures are required, use
directions given in the Training Guide.
2a. Gooch crucibles: Rinse with distilled water and
shake off excess. Crucible need not be completely
dry before re-use.
2b. Disc support (watch glass etc.): Rinse with dis-
tilled water. Dry completely before re-use.
2c'. If stronger cleaning measures are required, use
directions given in Training Guide.
V.J.If
(p. 27)
V.J ,2c
(p. 27)
K. Calculations
1. Use the following steps to
calculate total suspended
(non-filterable) solids,
mg/liter.
la. The calculation formula is:
Total suspended solids, mg/liter =
"gi^-r«!d^»m
-------
EFFLUENT MONITORING PROCEDURE: Total Suspended (Non-Filterable) Solids, mg/liter
Page No. 16-22
3PERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
K. Calculations (Conti
lued)
2.	Subtract the "weight of filter
(g)" on line 14 from the
"final weight of filter plus
residue (g)" on line 13.
3.	Write the difference on line
15 of the data sheet.
4.	Divide this difference on
line 15 by the "ml sample
filtered" on line 7 to get
a 7 decimal place answer.
5.	Write this answer on line 16.
2a. Example on data sheet:
line 13 - G.1413 g.
line 14 - 0.1293 q.
Difference = 0.0120 g.
2b. NOTE: This is the gram weight of the residue
which was on the filter disc.
2c. IMPORTANT: This gram weight of the residue (the
difference) should be greater than 0.0025 g. If
the weight of the residue (the difference) is less
than 0.0025 g, you should repeat the procedure and
filter a larger volume of the sample so more
residue is obtained.
3a. This has been done for the example in the third
"Sample" column.
4a. Example on data sheet:
line 15 = 0.01209 = 0.0001791 g/ml
line 7 67.0 ml
4b. NOTE: This is the gram weight of residue in each
ml of the sample.
5a. This has been done for the example in the third
"Sample" column
VII.K.2c.
fp. 30)
IX.K.3
(p. 32)
IX.K.5
(P. 32)
-------
EFFLUENT MONITORING PROCEDURE: Total Suspended (Non-Filterable) Solids, mg/liter
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
K. Calculations (Contln
jed)
6.	Multiply the 7 decimal place
answer on line 16 by 1,000,000
(Move the decimal point 6
places to the right).
7.	Write this answer on line 17.
8.	Round off answer on line 17 to
to the nearest whole mg,
9.	Write this answer on line 18.
6a. Example on data sheet:
line 16 is 0.0001791 g/ml x 1,000,000 = 179.1 mg/liter
6b. NOTE: This multiplication converts the gram weight
of residue per ml to the unit of mg/liter.
7a. This has been done for the example in the third
"Sample" column.
8a. Example on data sheet:
line 17, 179.1 mg/liter becomes: 179 mg/liter
9a. This has been done for the example in the third
"Sample" column.
9b. Records should be kept in a laboratory notebook.
IX.K.7
(P- 32)
U.K. 8a
(p. 26)
IX.X.9
(P. 32)
L. Reporting Data
1. Report total suspended
(non-filterable) solids,
mg/liter.
la. On any required record or report sheets.
IX.L.la
(F¦ 31}

1


Page No. 16-23
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EFFLUENT MONITORING PROCEDURE: Determination of Total Suspended (Non-Filterable)
Sol ids, mg/1iter
TRAINING GUIDE
SECTION	TOPIC
*1	Introduction
*11	Educational Concepts-Mathematics
III	Educational Concepts-Science
IV	Educational Concepts-Communications
*V	Field & Laboratory Equipment
VI	Field & Laboratory Reagents
*VII	Field & Laboratory Analysis
VIII	Safety
*IX	Records & Reports
*0nly these sections are used in this procedure.
Page No. 16-24
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EFFLUENT MONITORING PROCEDURE: Determination of Total Suspended (Non-Filterable)
Solids, mg/liter
INTRODUCTION
Section I
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
Suspended solids are insoluble solids that are in
suspension or dispersed in water, wastewater, or
other liquids. These are largely removable by
standard filtering procedures in a laboratory.
The term "suspended solids" is used here to refer
to the quantity of material removed from wastewater
under specified laboratory test conditions. The
test described in this instruction can be found in
the EPA Methods Manual on page 268, entitled "Resi-
due, Total Non-Filterable."
The amount of suspended solids in samples can be
used to indicate the efficiency of primary and
final settling tanks and the quality of plant
effluent. Thus the results of this test are used
for plant control and for regulatory requirements.
This procedure to determine suspended solids can
also be found in Standard Methods on page 94, en-
titled "Total Nonfiltrable Residue Dried at
103-105C (Total Suspended Matter)."
Glossary Water & Wastewater
Control Engineering. 1969.
WPCF, Washington, DC 20016
Methods for Chemical
Analysis of Water and
Wastes. 1974. EPA-MDQARL,
Cincinnati, OH 45268,
p. 268
Standard Methods for the
Examination of Water and
Wastewater. 14th ed.,
1976. APHA, Washington,
DC. p. 94
Paqe No. 16-25
-------
EFFLUENT MONITORING PROCEDURE: Determination of Total Suspended (Non-Filterable) Solids
mg/liter
EDUCATIONAL CONCEPTS-MATHEMATICS
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
1.15a -1
EXAMPLE of constant weights that differ only by a
possible balance error of-± O.OOOlg (0.1 mg;
True weight ='0.1286g
1st wt. obtained = 0.1287g (True + 0.1 mg)
2nd wt. obtained = 0.1285q (True - 0.1 mq)
Difference = 0.0002g
Thus to agree within possible balance error, the
difference between the two weights should not be more
than 0.0002g (0.2 mg).

1.15a.2
EXAMPLE of an acceptable difference between succes-
sive weights v/here the difference is not more than
O.OOOBg (0.5 mg):
1st wt. obtained = 0.1287g
2nd wt. obtained = 0.l283g
Difference = 0.0004g (0.4 mg)
Use the 2nd wt. obtained.

K.8a.
Rounding results to the nearest whole mg: If the
digit 0,1,2,3 or 4 is dropped, the preceding digit
is not altered.
EXAMPLE: 10.4 mg is rounded to 10 mg
Standard Methods for the
Examination of Water and
Wastewater. 14th ed., 1976
APHA, Washington, DC. p. 18

If the digit 5 is dropped, the preceding digit 1s
rounded off to the nearest even number.
EXAMPLES: 10.5 mg is rounded to 10 mg
11.5 mg 1s rounded to 12 mg
U.S. EPA, Handbook for
Analytical Quality Control
in Water and Wastewater
Laboratories. 1972, EPA-
AQCL, Cincinnati, OH 45268.
p. 7-2

If the digit 6,7,8 or 9 is dropped, the preceding
iiglt is Increased by one unit.


iXAMPLE: 10.6 mg 1s rounded to 11 mg

Page No. 16-26
-------
EFFLUENT MONITORING PROCEDURE:
Determination of Total Suspended (Non-Filterable) Solids
mg/liter
FIELD & LABORATORY EQUIPMENT
Section V
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
Gooch crucibles or filter disc supports (watch
glasses, etc.) should have identification marks which
will not be lost at the oven temperature of 103°-
105°C. Gooch crucibles with this type marking can
be purchased from laboratory supply companies. You
can permanently mark glass or porcelain surfaces with
an electrical marking tool or with marking ink
followed by firing in a flame. You can purchase the
tool or ink, or you can make marking solutions of
ferric chloride or of ordinary blue-black ink forti-
fied with a few grams of dissolved iron-potassium
tartrate. The marks are melted onto the surface by
firing in a flame or oven.
The suction flask does not require cleaning. Using
a soft brush, clean all other equipment with soap
or detergent. If stronger cleaning measures are
required, soak equipment in dilute acid or chromic
acid cleaning mixture.
After cleaning, rinse the equipment three times with
tap water and three times with distilled water.
The following do not have to be completely dry .
before using: Gooch crucibles, filter funnels,
filter holders, suction flasks.
The following should be completely dry before using:
graduated cylinders for measuring samples, filter
supports such as watch glasses.
Deslccants are hygroscopic materials capable of
absorbing moisture from air. Silica gel (SIO2) and
calcium sulfate (CaS04) are two commonly used
deslccants available from laboratory supply companies
These change color as they become saturated. The
moisture can be removed from the deslccant by heating
it in an oven.
Hamilton and Simpson, Quan-
titative Chemical Analysis.
1958. Macmillan, NY, NY
p. 40
U.S. EPA, Handbook for
Analytical Quality Control
in Water and Wastewater
Laboratories. 1972. EPA-
AQCL, Cincinnati, OH 45268,
p. 4-7
Paqe No. 16-27
-------
EFFLUENT MONITORING PROCEDURE: Determination of Total Suspended (Non-F1lterable) Sol Ids,
mg/liter
FIELD & LABORATORY ANALYSIS	Section VII
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
B.2a.
COLLECTION OF SAMPLES FOB THIS TEST:
Samples should be collected from a preagreed site
by a preagreed technique known to all parties con-
cerned. You should be familiar with the following
information since you record most of it on your
laboratory data sheet. You may be responsible for
actually collecting the sample; consult your super-
visor.




LOCATION -
Plant control and self-monitoring requirements will
be the basis for selecting places to collect samples.
Final collection points should be such that samples
drawn there are as representative of the entire
sample source as possible. Consult your supervisor.
Standard Methods for the
Examination of Water and
Wastewater. 14th ed.,
1976, APHA, Washington,
DC, p. 38

IDENTIFICATION -
Each collection location should be assigned a number
or simple identification code. Use this to label
samples from that location and to record on the lab
data sheet.




TYPE -
Permit requirements determine whether a grab or a
composite sample will be collected; consult your
supervisor. Mark type on sample container and on
laboratory data sheet.
Ibid.,
p. 40


TIME OF COLLECTION -
Mark time and date on sample container and on lab
data sheet.




CONTAINER -
The analyst should know what volume container 1s
required for each sample source. Containers should
be capped, of resistant (to adsorption of solids)
glass or plastic. Clean used containers by rinsing
with dilute hydrochloric acid solution, with tap
water (3 rinses) and with distilled water (3 rinses).
Shake out excess water.
Methods for Chemical Anal-
ysis of Water and Wastes.
1974. EPA-MDQARL, Cin-
cinnati, OH 45268, p. xi

COLLECTION -
Rinse container two or three times with sample, then
collect the sample. Consult the analyst about the
volume required from each sample source. Exclude
very large sol Ids like leaves, sticks, fish, lumps
of fecal matter, etc. Put cap on container.
Ibid.,
p. 268

Page No. 16-28
-------
EFFLUENT MONITORING PROCEDURE:
Determination of Total Suspended (Non-Filterable) Sol Ids,
mg/liter
FIELD & LABORATORY ANALYSIS
Section VII
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
SIGNATURE -
Sample Collector should sign name on container or
label so this Information can be recorded on the
lab data sheet.
STORAGE -
It 1s not practical to preserve and store these
samples. Analyze promptly to minimize chemical
and/or physical changes.
You want to filter a volume of sample such that pro-
longed drying times are not required (up to 0.200 g)
but that will yield a significant weight of residue
(at least 0.0025 q) on the filter disc.
Experience with samples from the same locations will
help you choose such volumes.
One useful guide (except for samples containing a
very high concentration of suspended matter, or
which filter very slowly) 1s to select a sample
volume of 14 ml or more per square cm of filter
area. (Recall that for a circle, area » 3.14 times-
radius squared.)
You can also use turbidity to estimate sample size.
If the sample has a turbidity of 50 units or less,
filter a liter of sample. For turbidity greater
than 50 units, filter sufficient sample to yield
up to 50 mg and not more than 100 mg of residue.
(If you are using a Gooch crucible, 50 mg is the
practical limit due to drying requirements.)
The time required for complete drying depends on
the amount and nature of the sol Ids on the filter
disc. The drying time given 1n this procedure Is
the MINIMUM time to be used.
1. If the sol Ids have a glassy, wet appearance
after the MINIMUM drying time, Increase this
drying time.
If you routinely run this test on samples from
the same source and check them for complete
drying (see I. In the procedure), you could
choose a smaller sample volume for future
determinations so that a longer drying time will
not be necessary.
Standard Methods for the
Examination of Water and
Wastewater. 13th ed.,
1971. APHA, New York, NY
p. 537
Ibid, p. 291
Page No. 16-29
-------
EFFLUENT MONITORING PROCEDURE: Determination of Total Suspended (Non-Filterable)
	 Solids, mg/liter
FIELD & LABORATORY ANALYSIS
Section VII
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
I.
K.2c.
The need to verify the complete drying of the
filter plus residue is important enough to warrant
the extra time required to make this check. The
weight of traces of water allowed to remain in the
residue would contribute significant error to the
final results in this test. The check for complete
drying presented in this section depends on ob-
taining a constant (same) weight after repeating
the heating, cooling and weighing cycle for the
filter plus residue.
If the weight of the residue is less than 0.0025g,
there is not enough weight to be significant for
this direct weighing method.
Methods for Chemical
Analysis of Water and
Wastes. 1974. EPA-
MDQARL, Cincinnati, OH
45268, p. 269
Standard Methods for the
Examination of Water and
Wastewater. 13th ed.,
1971. APHA, New York, NY.
p. 291
Page No. 16-30
-------
EFFLUENT MONITORING PROCEDURE: Determination of Total Suspended (Non-Filterable)
	 Solids, mg/liter
RECORDS & REPORTS
Section IX
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
B.2b.
B.2c.
L.la.
All laboratory records must be kept for three
years, preferably in a permanently bound notebook.
The time period is required by regulatory agencies.
Attached as the next page is a typical laboratory
data sheet for recording weights and for the
later calculation of final results for suspended
solids determinations.
Depending on your organizational set-up,	it may
be your job responsibility to enter this	data on
the plant operation record, state report	form,
etc. Check with your supervisor.
Page No. 16-31
-------
Typical Laboratory Data Sheet
for
TOTAL SUSPENDED (NON-FILTERABLE) SOLIDS, mg/llter
Name of Plant			
STEP
SUSPENDED SOLIDS
SAMPLE
SAMPLE
SAMPLE

B.2
Identification


INS #1
1
B.2
Type (grab, etc.)


GRAB
2
B.2
Date & Time Collected


5/1/74 0900
3
B.2
Sample Collector


Tom Sampler
4
C.4
Filter Identification


WG2
5
E. 1
Date & Time Analysis began


5/1/74 1100
6
E.8
ml Sample Filtered


67.0
7
H.6
1st weight of Filter*
plus Residue (g)


0.1426
8
1.11
2nd weight of Filter*
plus Residue (g)


0.1416
9
1.14
Difference (lst-2nd)


0.0010

1.15
3rd weight of Filter*
plus Residue (g)


0.1413
11
1.15
Difference (2nd-3rd)


0.0003
12
1.15
Final weight of Filter*
plus Residue (g)


0.1413
13
C.7
Weight of Filter* (g)


0.1293
14
K.3
Find Difference (g) by subtracting
Line 14 from Line 13


0.0120
IS
K.5
Divide to 7 decimal places:
(line 15) difference (q)
(line 7) ml sample filtered


0.0001791
16
K.7
Multiply Line 16 by 1000 000
(move decimal point 6 places Rt.)


179.1
17
K.9
Round answer on Line 17
to nearest whole number


179 mg/1
18
1.16
Analyst


Mary Analyst
19
*"Filter" means the filter disc 1f a funnel type filtration assembly 1s used. If Gooch
crucibles are used "filter" means the crucible containing a filter disc.
Page No. 16-32
-------
A PROTOTYPE FOR DEVELOPMENT OF
ROUTINE OPERATIONAL PROCEDURES
for
SETTLEABLE SOLIDS, ml/liter
(IMHOFF SETTLING CONE)
as applied in
WASTEWATER TREATMENT FACILITIES
and in the
MONITORING OF EFFLUENT WASTEWATERS
Developed by the
National Training and Operational Technology Center
Municipal Operations and Training Division
Office of Water Program Operations
U.S. Environmental Protection Agency
CH.SOL.set.EMP.l.5.77
Page No. 17-1
-------
EFFLUENT MONITORING PROCEDURE: Settleable Solids, ml/Hter (Imhoff Settling Cone)
This operational procedure was developed by:
NAME	Audrey D. Kroner
ADDRESS EPA, OWPO, NTOTC, Cincinnati, Ohio 45268
POSITION Chemist-Instructor
EDUCATION AND TECHNICAL BACKGROUND
B.A. - Edged iff College
1 year Industrial Research Chemist
8 years Secondary School Chemistry Instructor
4 years DHEW-DI Water Quality Program Chemist
7 1/2 years DI-EPA Chemist-Instructor
Page No. 17-3
-------
EFFLUENT MONITORING PROCEDURE: Settleable Solids, ml/liter (Imhoff Settling Cone)
1.	Analysis Objective:
To determine settleable matter on a volume (ml/1) basis.
2.	Brief Description of Procedure:
A one liter sample is poured into an Imhoff Cone and the volume (ml/I)
of settleable solids is recorded after a one hour settling period.
3.	Applicability of this Procedure:
a.	Range of Concentration:
The one Source of Procedure* cited in the Federal Register Guidelines does
not state a range of concentration. This EMP includes a procedure for
cases when the settleable solids exceed the graduations on an Imhoff Cone.
b.	Pretreatment of Samples:
The Federal Register Guidelines do not specify any pretreatment, nor does
the Source of Procedure*.
c.	Treatment of Interferences in Samples:
The Source of Procedure does not note any interferences to this deter-
mination.
*Source of Procedure: Standard Methods for the Examination of Water and Waste
water, 14th ed., APHA, Washington, D.C., p. 95.
-------
EFFLUENT MONITORING PROCEDURE: Settleable Solids, ml/liter (Imhoff Settling Cone)
Flow Sheet for Determination:
1.	Mix sample
2.	Fill cone
3.	Settle 45 minutes
4.	Stir gently or swirl
5.	Settle 15 minutes
6.	Read results
Page No. 17-5
-------
EFFLUENT MONITORING PROCEDURE: Settleable Solids, mg/liter (Imhoff Settling Cone)
Equipment and Supply Requirements:
A.	Capital Equipment: None
B.	Reusable Supplies:
1.	Imhoff Settling Cone, glass or plastic, with or without stopcock, graduated
to 40 ml and with a graduation at 1 liter volume
2.	Imhoff Cone support, 3 place
3.	Imhoff Cone brush (or centrifuge tube brush)
4.	Stirring rod, the same length as cone
5.	Timer, interval, 60 minute minimum with alarm
C. Consumable: None
Page No. 17-6
-------
EFFLUENT MONITORING PROCEDURE; Settleable Solids, ml/liter (Imhoff Settling Cone)
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
SETTLEABLE SOLIDS
A. Preparation for the
Determination
1.	Gather equipment for determi-
nation.
2.	Clean the cone and rod.
3.	Bring sample to work area.
4.	Record sample identification.
5.	Record cone Identification
Information.
la. Imhoff cone
lb. Imhoff cone support
1c. Long stirring rod
Id. Timer
2a. If required
2b. Water drains without leaving many droplets.
3a. Sample should be at hand before continuing with
this test.
4a. In laboratory notebook
4b. In column to be used for sample
4c. Identification, type, date and time collected,
sample collector.
5a. If more than one cone is to be in use
5b. Cones can be numbered with a lab marking pen or
pencil.
5c. Record identification in laboratory notebook
5d. In column-for corresponding sample
5e. Labeled "Cone Identification"
I
(P. 11)
V.A.2b.
(P. 12)
VII. A.3a.
(P- 13)
IX.A.4a.
(P. 15)
IX.A.4c.
(P. 15)
IX.A.5.
(P- 16)
B. Determination
1. Record date and time.
la. In laboratory notebook
lb. In column to be used for sample
lc. Labeled "Date & Time Analysis Began"
IX.B.l.
(P- 16)
Page No. 17-7
-------
EFFjUENT MONITORING PROCEDURE: Settleable Solids, ml/liter (Imhoff Settling Cone)
Page No. 17-8
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
B. Determination
(Continued)
2.	Thoroughly mix the sample.
3.	Fill the Imhoff Cone to the
1 liter mark with sample.
4.	Set timer at 45 min.
5.	Allow the sample to settle 45
minutes,
6.	Gently stir the sample along
the inner wall of the cone,
7.	Set timer at 15 minutes.
8.	Allow the sample to settle for
15 more minutes.
2a. Use a stirrina rod or swirl gently.
2b. Do not shake the sample.
5a. in quiescent location
5b. not in sunlight
6a. to dislodge solids clinging to inner wall.
6b. Use a long stirring rod or else spin the cone.
VII.B.2b.
(p. 14)
VII.B.5a.
(P- 14)
VII.B.5b.
(P- 14)
C. Results
1.	Read the final volume of
settled solids,
2.	Record the final volume of
settled Solids in the cone
la. in ml
lb. with eye at level of surface of settled matter.
1c. In rare cases, the final volume of settled matter
may be above the 40 ml mark
2a. in laboratory notebook,
2b. in column used for that sample,
2c. labeled "Final Volume of Settleable Solids,
ml/liter",
2d. to nearest whole ml per liter.
VII.C.lc.
(p: i4)
IX.C.2a.
(P. 15)
IX.C.2.
(p. 16)
-------
EFFLUENT MONITORING PROCEDURE; Settleable Solids, ml/liter (Imhoff Settling Cone)
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
C. Results(Continued)
3.	Sign the laboratory data sheet.
4.	Report Settleable Solids,
ml/1 iter
3a. Labeled "Analyst"
4a. on any required record sheets.
IX.C.3.
(P. 16)
IX.C.4a.
(P. 15)
D. Cleaning Equipment
1.	Discard cone contents.
2.	Rinse the cone and stirring
rod
3.	Complete cleaning the cone
and rod as soon as possible,
2a. with tap water
3a. to prevent algal growth
V.D.2a.
(P. 12)
V.D.3a.
(P. 12)




Page No. 17-9
-------
TRAINING GUIDE
SECTION	TOPIC
*1	Introduction
II	Educational Concepts-Mathematics
III	Educational Concepts-Science
IV	Educational Concepts-Communications
*V	Field & Laboratory Equipment
VI	Field & Laboratory Reagents
*VII	Field & Laboratory Analyses
VIII	Safety
*IX	Records & Reports
*0nly these sections are used in this procedure.
Page No. 17-10
-------
EFFLUENT MONITORING PROCEDURE: Settleable Solids, ml/liter (Imhoff Settling Cone)
INTRODUCTION

TRAINING GUIDE NOTE
REFERENCES/RESOURCES

Settleable solids would be that matter in wastewater
which will not stay in suspension during a pre-
selected settling period (such as one hour) but
either settles to the bottom or floats to the top.
In the Imhoff Cone test, the settling period is one
hour and the quantity of solids is expressed by
volume (ml/1) of settled matter.
The test described in this instruction is Method
208F, page 95 in Standard Methods.
This test is used as a Control Check on the proper
functioning of treatment processes. Usually, samples
are drawn from the raw waste influent and from
effluents of the primary and secondary processes.
By comparing the quantity of settleable solids among
these samples, the effectiveness of removing solids
(and turbidity) can be determined. Although the
test is not quantitative, it is very useful as a
process control test to indicate the volume of sludge
which must be withdrawn from a particular process.
This test is listed in the Federal Register "Guide-
lines Establishing Test Procedures for the Analysis
of Pollutants." The only reference cited for the
procedure is Standard Methods, 14th ed., page 95.
Glossary Water and Waste-
water Control Engineering.
1969. WPCF, Washington,
DC 20016.
Standard Methods for the
Examination of Water and
Wastewater. 14th ed.,
1975. APHA, Washington,
DC 20036.
Richmond, M.S., et.al.
Simplified Laboratory
Procedures for Wastewater
Examination. Pub. No. 18,
1968. WPCF, Washington,
DC 20016.
Page No. 17-11
-------
EFFLUENT MONITORING PROCEDURE: Settleable Solids, ml/liter (Imhoff Settling Cone)
SECTION V
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
A.2b.
D.2a,
D.3a.
Using an Imhoff cone brusli, clean cones with soap or
detergent water. Rinse with tap water, shake and
invert to drain dry. A few droplets of water may
remain on the walls of the cone, especially if
plastic cones are used. This is permissible. If
stronger cleaning measures are required, rinse cone
with dilute acid or chromic acid cleaning mixture,
rinsewith tap water and drain. (Chromic acid
cleaning mixture contains, sulfuric acid and should
be used carefully. Sulfuric acid causes severe
burns.)
It is always good practice to rinse equipment with
tap water as soon as possible after use to facilitate
cleaning.
Use cleaning procedure described above in A.2b.
Handbook for Analytical
Quality Control in Water
and Wastewater Laboratories.
1972. U.S. EPA, NERC,
Cincinnati, OH 45268
Ibid,
Page No. 17-12
-------
EFFLUENT MONITORING PROCEDURE: Settleable Solids, ml/liter (Irahoff Settling Cone)
SECTION VII
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
A.3a.
COLLECTION OF SAMPLES FOR THIS TEST:
You should be familiar with the following information
since you record most of it on your laboratory data
sheet. You may be responsible for actually collect-
ing the sample; consult your supervisor.
Location -
Plant control and self'-monitoring requirements will
be the basis for selecting places to collect samples.
Final collection points should be such that samples
drawn there are as representative of the entire
sample source as possible. Consult your supervisor.
Identification -
Each collection should be assigned a number or simple
identification code. Use this to label samples from
that location and to record on the lab data sheet.
Type -
Collect a grab sample immediately before the test is
to be started. Mark type (grab) on sample container
and on lab data sheet.
Time of Collection -
Mark date and time on sample container and on lab
data sheet.
Container -
1000+ ml volume, capped, resistant (to adsorption
of solids) glass or plastic. Clean used containers
by rinsing with dilute hydrochloric acid solution,
with tap water (three rinses) and with distilled
water (three rinses). Shake out excess water.
Collection -
Rinse container two or three times with sample, then
collect about 1000 ml of sample. Exclude very
large solids like leaves, sticks, fish, .lumps of
fecal matter, etc. Put cap on container.
Signature -
Sample Collector should sign name on container or
label so this information can be recorded on the lab
data sheet.
Storage-
It is not practical to preserve and store these
samples. Analyze promptly to minimize chemical and/
or physical' .changes.
Standard Methods for the
Examination of Water and
Wastewater. 14th ed., 1975.
APHA, Washington, D.C.
20036, D. 38
Ibid.
Ibid, p. 91
Page No. 17-13
-------
EFFLUENT MONITORING PROCEDURE: Settleable Solids, ml/liter (Imhoff Settling Cone)
SECTION VII
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
B.2b.
B.5a.
B.	5b.
C.lc,
Page No. 17-
Entrained air interferes with settling.
Vibrations interfere with settling.
Sunlight can cause heat currents in the sample which
interfere with settligg.
It is possible to have a final volume of settled
matter greater than the 40 ml (largest) volume
marking on the cone. If this happens:
1.	Use a grease or wax pencil and put a mark on the
outside of the cone at the level of the surface
of the settled matter.
2.	Discard the sample from the cone.
3.	Using a 100 ml graduated cylinder and tap water,
fill the cone up to the mark you made on the
cone. Keep a count of the volumes of water you
add.
4.	The total ml of water added to reach the mark
you made o'n the cone are the ml of settled
matter you should report as the test result.
-------
EFFLUENT MONITORING PROCEDURE: Settleable Solids, ml/lfter (Imhoff Settling Cone)
SECTION IX
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
A.4a.
C.2a.
A.4c.
C.4a.
All laboratory records should be kept 1n a per-
manently bound book. Tlrts 1s especially Important
as documentation for any future questions about
data required by regulatory agencies.
Attached as the next .page is a typical laboratory
data sheet for recording information about this
analysis.
Depending on your organizational set-up, it may be
your responsibility to enter the result on the
plant operation record, state report form, NPDES
report form, etc.. Check with your supervisor.
Handbook for Analytical
Quality Control in
Water and Wastewater
Laboratories. 1972. U.S.
EPA, NERC, Cincinnati,
OH 45268
I
Page No. 17-15
-------
Typical Laboratory Data Sheet
for
SETTLEABLE SOLIDS, ml/liter
(Imhoff Settling Cone)
Name of Plant
STEP
SETTLEABLE SOLIDS
SAMPLE
SAMPLE
SAMPLE
A.4.
Identification


IN n
A.4.
Type (Grab, etc.)


GRAB
A.4.
Date & Time Collected


9/23/74 9:55
A.4.
Sample Collector


John Sampler
A.5.
Cone Identification


#1
BJ.
Date & Time Analysis began


9/23/74 10:00
C.2.
Final Volume of Settleable
Solids, ml/liter


26 ml/liter
C.3.
Analyst


Tom Analyst





Page No. 17-16
-------
A PROTOTYPE FOR DEVELOPMENT OF
ROUTINE OPERATIONAL PROCEDURES
for the
REPORTING OF SELF - MONITORING DATA
as applied 1n
WASTEWATER TREATMENT FACILITIES
and in the
MONITORING OF EFFLUENT WASTEWATERS
Developed by the
National Training and Operational Technology Center
Municipal Operations and Training Division
Office of Water Program Operations
U.S. Environmental Protection Agency
-------
EFFLUENT MONITORING PROCEDURE: REPORTING OF SELF-MONITORING DATA
This Procedure was developed by:
NAME Charles E. Sponagle
ADDRESS EPA, OWPO, f!TOTC, Cincinnati, Ohio 45268
POSITION Sanitary Engineer-Instructor
EDUCATION AIID TECHNICAL BACKGROUND
B.C.E. - Manhattan College, 1943
M.S. in C.E. - University of Minnesota, 1948
Professional Registration: State of New York
With Federal Water Pollution Control Program since 1948, with
various assignments at Program Headquarters, Regional Offices,
and Field Stations, including positions as
Staff Engineer, then Chief, Water Quality Section
Denver Regional Office
Staff Engineer, then Regional Construction Grants Program
Director, Denver Regional Office
Regional Construction Grants Program Director,
Cincinnati Regional Office
Director, Colorado River Basin Water Quality Control Project,
Denver Colorado
Industrial Wastes Consultant, Technical Advisory and
Investigations Branch, Cincinnati, Ohio
Participation in and Direction of numerous in-plant industrial
waste surveys and stream studies in New York, Colorado,
New Mexico, Main?, Utah
With National Training Center, September 1969 to date.
Page No. 18-3
-------
EFFLUENT MONITORING PROCEDURE: Reporting of Self-Monitoring Data
1.	Objective: To enable the student to complete the NPDES Discharge
Monitoring Report, EPA Form T-40^4-74), or EPA Form 3320-1(10-/2).
2.	Description of Procedure:
Self-monitoring data obtained by a permit holder under the terms of his
permit must be reported to the regulatory agency periodically, using the
proper NPDES reporting form. The manner in which such data should be
reported on EPA Form T-40 is illustrated in this procedure. Additional
information required to complete the form is also indicated.
Assumed conditions used to illustrate completion of the form are:
1.	Reporting of data on a monthly basis is required.
2.	Self-monitoring data developed over a period of one
month is as shown in Table I, Page 5
3.	Effluent limitations specified in the permit are as
shown on Table II, Page 6
4.	Monitoring requirements specified in the permit are
as shown in Table II, Page 6
5.	All required data has been obtained in accordance
with permit requirements.
Page No. 18-4
-------
EFFLUENT MONITORING PROCEDURE: Reporting of Self-Monitoring Data
TABLE I
SELF - MONITORING DATA
September 1974

SEWAGE FLOW
RAW
INFLUENT

FINAL
EFFLUENT

Date
Treated
BOD,
T.S.S.
BOD
T.S.S.
Fecal
PH


0

3

Coliform

9Pd
mg/1
mg/1
mg/1
mg/1
N/100 ml

1
720,100





7.4
2
609,000





7.5
3
326,900
170
171
16
12
350
7.6
4
367,500





7.4
5
323,900





7.5
6
458,500
160
168
15
16
540
7.7
7
571 ,000





5.4
8
508,600





7.6
9
146,000
200
200
20
25
180
7.9
10
253,000





7.2
11
406,800





7.1
12
519,200
190
198
20
25
170
7.6
13
328,600





7.5
14
413,100





7.6
15
699,000





8.0
16
708,900
150
180
35
60
220
8.0
17
806,700





9.2
18
714,800





8.0
19
169,100





9.1
20
272,900
170
170
19
19
240
7.5
21
713,200





7.8
22
671,900





7.0
23
761,800
150
186
20
23
110
7.4
24
642,900





7.5
25
314,900





7.4
26
291,600
190
195
20
20
130
7.5
27
240,700





7.4
28
478,900





7.4
29
525,600
190
195
25
25
280
7.6
30
670,100





7.8
Total
14,635,200






Average
487,800






Page No. 18-5
-------
Paqe No. 18-6
EFFLUENT MONITORING PROCEDURE: Reporting of Self-Monitoring Data
TABLE II
EFFLUENT LIMITATIONS AND MONITORING REQUIREMENTS
DISCHARGE LIMITATIONS
EFFLUENT CHARACTERISTICS
Biochemical Oxygen Demand (5-day)
Suspended Solids
pH - standard units
Fecal Coliform - organisms/100 ml
Flow - mgd
Concentration
kg/day
MINIMUM
*30
*30
5.0-S
200
Weekly
Monthly Weekly
Measurement
Sample
Average
Average Average
Frequency
Type
45
70
twice weekly
24 hr. composite

(150)

45
80
twice weekly
24 hr. composite

(180)
(not to
be averaged)
twice weekly
grab
400
— —
twice weekly
grab
daily
recording
* The arithmetic mean of the values for effluent samples measuring biochemical oxygen demand
(5-day) and suspended solids collected in a period of 30 consecutive days shall not exceed
15 percent of the arithmetic mean of the values for influent samples collected at approximately
the same times during the same period (85 percent removal—minimum).
* Whichever is the more stringent.
-------
EFFLUENT MONITORING PROCEDURE: REPORTING OF SELF-MONITORING DATA
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
A.
Description of
EPA Forms


I.A
(P. 27)
B.
Identification
of Permit Holder
and Discharge
1.
Enter Name and Address
of Permit Holder
la.
lb.
In space provided at left of Instructions
May already be entered by permit-issuing
authority.



2.
Enter State in block
labelled "ST"
2a.
2b.
Use standard two-letter postal code
See notes la and lb above.



3.
Enter Permit number in
block for same.
3a.
See notes la and lb above.



4.
Enter discharge number
in "Dis" block.
4a.
4b.
As identified in permit (001, 002, etc.)
See notes la and lb above.



5.
Enter Discharge code
5a.
5b.
For municipal wastewater discharges, the
number is 4952.
See notes la and lb above.



6.
Enter Latitude and
Longitude of discharge.
6a.
6b.
If known.
See notes la and lb above.



7.
Enter reporting period in
appropraite blocks.
7a.
7b.
7c.
In this procedure the 30-day period for the
month of September is used.
Will be specified in permit.
See notes la and lb above.




This portion of the report form, completed in accordance with assumed permit conditions, and
the data of Table I, is shown in Fig. 1:
Page No. 18-7
-------
fFFLUENT MONITORING PROCEDURE: REPORTING OF SELF-MONITORING DATA
Page No. 18-8



TRAINING
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
GUIDE NOTES
NATIONAL POLLUTANT DISCHARGE ELIMINATION SYSTEM
DISCHARGE MONITORING REPORT
Form Approved
OMB NO. 158-190073
r	i
City of Noname, Dept. of Environmental Services
184 Any Street
Noname, Anystate, 12345
L
J
AN.
NM?MSfi7
PERMIT NUMOEft
QQ1
4952
47°20'4"
98°26'11"
LATITUDE
LONGITUOC
REPORTING PERIOO FROM
>• j II m-2> 124-211
HS-27> !»•«> HQ-Si)
714
019
oil
TO
714
019
310
YEAR
MO
OAV

YEAR
wo
DAY
INSTRUCTIONS
1.	Provide dates (or period covered by this report in spaces merited "REPORTING PERIOD"
2.	£ot«r reported miniata, «v«r«(e and maximum values under "QUANTITY" end "CONCENTRATION"
is fee units specified for each parameter as appropriate. Do not enter values in boxes containing
asterisks. "AVERAGE" is average computed over actus! tine discharge it operating. "MAXIMUM"
and "MINIMUM" ere extreme values observed during the reporting period.
1 Specify the amber of analyzed samples that exceed the natintuB (end/or minitmaa •» epproprimte)
pemit conditions in the columns labeled "No. Ek." If nont, enter "O".
4.	Specify frequency of analysis for each parameter as No. analyses/No. days, (e.g , "3'7" is oquivm-
ient to 3 melytet performed every 7 deyt) If continuous enter "CONT."
5.	Specify sample typ*("grab" or "	hr. composite") as ^plicable. If frequency was continuous,
eater "NA"-
6.	Appiophite signature is required on bottom of this font.
7.	Remove carbon and retain copy for your records.
8- Fold along dotted lines, staple sod mail Original to office specified in permit.
:in. 1
C. Flow Data.
1. "Quantity"
section.
1.	Enter minimum and maximum
flows during the reporting
period in the correspond-
ing spaces on the
"reported" line.
2.	Enter Average flow during
reporting period in
corresponding space on
"reported" line.
3.	Enter on "Permit Condition"
line the average and
maximum daily flows
specified in the permit.
la.
lb.
2a.
2b.
3a.
3b.
Minimum flow of 0.15 MGD on Sept. 9. (Table I)
Maximum flow of 0.81 MGD on Sept. 17.(Table I)
Add the flows reported during the reporting
period. Divide this total by the number of
flows reported.	g35
From Table I, Average =	= 0.49 MGD
If unspecified in Permit, place a dash in the
appropriate space or spaces.
May already have been entered by permit-issuing
authority.
IX.C.1.1
(p- 35)
IX.C.I.3.3a
(p. 35)
-------
efflue;;t	procedure: reporting of self-monitoring data
STEP SEQUENCE
I ?: TORi'iAT i ori / O P E RAT I n g goals/specifications
TRAINING
GUIDE NOTES
Flow Data (cont.)
2. "Concentration1
section
3. "Frequency of
Analysis"
Column
4. "Sample Type"
Column
4. On the "Reported" line,
in the "No. Ex" space,
enter the number of times
during the reporting
period that the maximum
daily flow specified in
the permit was exceeded
1. Place dashes in the
"Units"space and in the
"No. Ex" spaces on the
"Reported" line.
1.	On the "Reported" line
enter the frequency with
which flows were measured
during the reporting
period.
2.	On the "Permit Condition"
line enter the frequency
of flow measurement as
specified in the permit.
1. Enter Dashes on both lines
4a. If none, enter "0".
4b. If a maximum daily flow is not specified in
the permit, place a dash in this space.
la. May already be entered by the Permit-issuing
authority.
la. Daily, Weekly, Continuous (Cont.), etc.
2a. May already be entered by Permit-issuing
authority.
This portion of the report form, completed in accordance with assumed permit conditions, and the data of
Table I, is shown in Fig. 2:
Paae No. 18-9
-------
EFFLUENT MONITORING PROCEDURE: REPORTING OF SELF-MONITORING DATA
Paae No. 18-10



11'A IMI N(j
OPERATING PROCEDURES
STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
GUILE NOTES

parameter

< 3 cmrd only)
IK-4S'
quantity
i S4-6 11

«2«J
(4 cmtd only)
CONCENTRATION
«e • 5 3' t *• 6 •


T
FREQUENCY |
I
SAMPLE !


MINIMUM
AVERAGE
MAXIMUM
UNITS
NO
EX
MINIMUM
!
AVERAGE MAXIMUM
UNITS
| NO
' EX
ANALYSIS !
1
TYPE j

FLOW
REPORTED
0.15
0.49
0.81
MHO
_
******

	
1
	1	J
Cont. 1 - 1

PC RMI T
CONDITION

-


¦
......


i
Cont.
j
ric|. 2
pH Data
1. "Quantity"
section
1. On the "Reported" line
enter the minimum and
maximum pH values occurring
during the reporting
period.
2.	On the "Permit Condition"
line enter the minimum and
maximum pH values spec-
ified in the permit.
3.	On the "Reported" line,
in the "Mo. Ex" space,
enter the total number of
times that the pH exceeded
the maximum allowed by the
permit, and was less than
the minimum allowed by the
permit.
la. Although the permit requires that pH be
determined twice weekly, a pH was run each day
during the month. The report form must be
prepared on the basis of all 30 results,
lb. Minimum pH 5.4 on Sept. 7 (Table I)
1c. Maximum pH 9.2 on Sept. 17. (Table I)
2a. Permit requires pH to be between 6.0 and 9.0
at all times (Tab!e II)
2b. May already be entered on the form by the
permit-issuing authority.
3a. The maximum permit requirement of 9.0 was
exceeded twice - once on Sept. 17, and again
on Sept. 19. (Table I)
3b. The pH was less than the minimum permit
requirement of 6.0 on Sept. 7. (Table I)
-------
EFFLUENT t-'ONITO^ItiS PROCEDURE: REPORTING OF SELF-MONITORING DATA
1
PPCCiyJSES | STEP SEQUENCE
INFORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
1 f
1
D. pH Data (cont.) i
t
2.	"Concentration" 1. Place dashes in the
section "Units" space and in the
"No. Ex" space on the
"Reported" line.
3.	"Frequency of 1. Enter frequency of
Analysis" analysis during the
column. reporting period on the
"Reported" Tine.
2. Enter required frequency
of analysis as specified
in permit on the "Permit
Condition" line.
4.	"Sample Type" 1. On the "Reported" line
column. indicate the type of
sample on which the
analysis was performed.
2. On the "Permit Condition"
line enter the type of
sample specified by the
j permit.
t
1
1
3c. Permit requirements were violated a total of
3 times during the reporting period. A "3"
should be entered on the Form.
3d. If the pH had riot exceeded the permit limits,
a "0" would be entered.
la. May already be entered by Permit-issuing
authority.
la. The actual frequency of analysis is reported.
lb. pH was run each day. The frequency of analysis
is reported as 7/7, which indicates that 7
analyses were performed every seven days.
2a. The permit requires that pH be run twice weekly.
(Table II) A 2/7 is entered on this line,
which indicates that 2analyses were to be
performed every 7 days.
2b. May already be entered by the Permit-issuing
authority.
la. A grab sample is assumed here. "Grab" is
entered.
2a. A grab sample is specified. (Table II) Enter
"Grab".
2b. May already be entered by the Permit-issuing
authority.

Page No. 18-11
-------
Page No. 18-12
P"0CEDl:RE: REPORTING OF SELF-MONITORING DATA



TRAINING

STEP SEQUENCE
ImFORMATION/0PERATING GOALS/SPECIFICATIONS
GUIDE NOTES
D. pH Data (cont.)
This portion of the report form, completed in accordance with assumed permit conditions, and the data of
Table I, is shown in Fig. 3 below:

PARAMETER

(3 card only)
'*• «5i
quantity
1
: 9»« 1'

162-03
(4 csrd on lyj
> 3I-45I
CONCENTRATION
M • 5 3 5 4-6'

FREQUENCY
SAMPLE !


MINIMUM
AVERAGE

MAXIMUM
| UNITS
1 NO
1 "
MINIMUM
AVERAGE MAXIMUM
¦ NO 1
UNITS Ex 1
OF
ANALYSIS
TYPE j





******


fcT/VNDARD







PH


5.4

9.2
U.


_ '
7/7
Grab


PEHMi T
CONDITION
6.0
******

9.0

I
******

~ ¦
[
ro
Grab
Fin. 1
E. BODg Data, Final





Effluent.





1. Computation of
Quantities
1. For each reported
analytical result,
calculate the quantity of
B0D5 discharged in Kg/day.



II.E.l
(P- 30
2. "Quantity"
section
1. On the "Reported" line
enter the minimum, average,
and maximum quantities
discharged over the
reporting period, in the
appropriate spaces.
la.
lb.
lc.
Minimum
Average
Maximum
-	11.0 Kg/day
-37.5 Kg/day
-	94 Kg/day

-------
£,) i _ l_- - < ' i ;'.I>1 I	* ivLv.JUnu •
REPORTING OF SELF-MONITORING DATA
r ~ •.-r - • • ^ crn^T-'
L ' L.\1 X.i-J •
STEP SEQUENCE
^FORMATION/OPERATING GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
E. BODg Data, Final
Effluent.(cont.)
3. "Concentration"
section.
j 3.
On the "Permit Condition"
line enter the average and
maximum quantities
permitted to be discharged
1.
2.
In the ".No. Ex" column,
on the "Reported" line,
enter the number of times
that the maximum daily
discharge of BOD5 (Kg/day)
allowed by the permit has
been exceeded during the
reporting period.
On the "Reported" line
enter the minimum, aver-
age, and maximum BOD5
concentrations observed
during the reporting
period.
On the "Permit Condition"
line enter the average and
maximum concentrations
specified in the permit.
2a. Average BOD5 discharge over a 30-consecutive-day
period is specified as 70 Kg/day. (Table II)
2b. A Maximum daily discharge limitation is not
indicated in the permit. (Table II) A dash
is therefore placed in the "maximum" space.
2c. May already be entered by the Permit-issuing
authority.
3a. If none, enter a "0".
3b. Since no maximum daily discharge limitation is
specified in the permit (Table II), a dash is
placed in this space.
la. Minimum - 15 mg/1
lb. Average - 21 mq/1
lc. Maximum - 35 mg/1
2a. Average BOD5 concentration over a 30-
consecutive-day period is specified as 30 mg/1.
(Table II)
2b. A maximum concentration is not indicated in the
permit conditions (Table II). Therefore place
a dash in this space.
2c. May already be entered by the Permit-issuing
authority.
II.E.I
(p- 30)
Page No. 18-13
-------
E~L-;~:7 i'OMIT-j^T^GPSCCEDVt-E: REPORTING OF SELF-MON ITORIfJG DATA
Page No. 18-14
C=£VTI\3 PirCEDlv.ES | STEP SEQUENCE
I i IF CRHATI C'< / OP E RAT I iS COALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
1
E. BODg Data, Final 3. In the "No. Ex" column,
\ the "Reported" line, enter
Effluent, (cont.) thg Qf times thflt
the maximum concentration
allowed by the permit has
j been exceeded during the
reporting period.
4.	"Frequency of 1. On the "Reported" line
Analysis" enter frequency of
column ! analysis during the
j reporting period.
i
! 2. On the "Permit Condition"
j line enter required
| frequency of analysis as
! specified in permit.
1
5.	"Sample Type" 1. On the "Reported" line
column enter the type of sample
on which the analysis was
performed.
2. On the ''Permit Condition"
line enter the type of
sample specified in the
permit.
This portion of the Report Form, completed in accoi
Table I, is shown in Fig. 4:
3a. If none, enter a "0".
3b. Since no maximum concentration is specified in
the permit (Table II) enter a dash in this
space.
la. Enter 2/7, indicating that the analysis was
performed twice weekly {Table I).
2a. Permit requires analysis twice weekly
(Table II). Enter 2/7.
2b. May already be entered by the Permit-issuing
authority.
la. Enter "24-Hr. comp." since it is assumed in
this procedure that the data has been obtained
in accordance with permit requirements.
2a. Enter "24-Hr. comp." (Table II).
2b. May already be entered by the Permit-issuing
authority.
rdance with assumed permit conditions, and the data of

-------
crf.UcuT r.Cv'lTORL^PRCCECuRE: REPORTING OF SELF-MONITORING DATA


i

1 TRAINING
C" ' T

j STEP SEQUENCE
INFCRMATIOii/OPERATING GOALS/SPECIFICATIONS
j GUIDE NOTES

	
	
(J cmrd only)

II.F.I
(p. 31)
Page No. 18-15
-------
Page No. 13-16
3. "Concentration"
section

"ior;/ope:mir;g goals/specifications
F. Percent Removal	j
BODc (cont.)	;•
3	I
2. "Quantity"	I
section	i
3.
1.
On the "Reported" line
enter the minimum,
average,and maximum
percent removals in the
appropriate spaces.
On the "Permit Condition"
line enter the minimum
and average removals
required by the permit.
la.
lb.
1 c.
2a.
j 2b.
I 2c.
In the "No. Ex" column,
on the "Reported" line,
enter the number of times
that the minimum percent
removal required in the
permit was not obtained.
Enter dashes in the
"Units" space and in the
"No. Ex" space on the
"Reported" line.
3a.
la.
This portion of the Report Form, completed in accordance
Table I, is shown in Fig. 5:
Minimum - 77%
Average - 87.9%
Maximum - 91Z
There is no minimum removal requirement in the
assumed permit conditions (Table II). Enter
a dash in this space.
Average removal required over a 30-consecutive-
day period is 85%. (Table II).
May already be entered by the Permit-issuing
authority.
There is no minimum removal requirement in the
assumed permit conditions (Table II). Enter
a dash in this space.
May already be entered by the Permit-issuing
authori ty.
with assumed permit conditions, and the data of
TRAINING
GUIDE NOTES
-------
"rFL'.'ET :-*C:iITC~X:.5G P^CLEC'jRE: reporting of self-monitoring data

s
1
1

*
!

TRAINING
. . u\ i - • >.
~ r*¦*<	,—'Tc !
• ' • J '-S t
S7E? SEQUENCE
} iNFrn?-iAi
"ICn/OPERATING GGALS/SPECIFICATIONS
GUIDE NOTES
¦ ii- r								¦»«-»~>	i >> -701

PARAMITCft

(3 emrtamtrf

t

l«3-«9)
(4 cird only)
CONCENTRATION
(«•«»! 9*-«>i


FREQUENCY
OF
ANALYSIS
SAMPLE


MINIMUM
AVERAGE
MAXIMUM
UNITS
MO.
EX
MINIMUM
AVERAGE
MAXIMUM
UNITS
NO.
EX
TYPE

PERCENT REMOVAL
ROORTID
77
87.9
91
'
-
******
******
******

.



BOD
s
P (Mil T
CONDITION
-
85
******
¦



"
¦


Fig. 5
G. Suspended Solids,
Final Effluent
1. Computation
of Quantities
2. "Quantity"
section
1. For each reported

II.G.I
analytical result,

(P- 32)
calculate the quantity of

suspended solids


discharged in Kg/day.


1. On the "Reported" line
la. Minimum - 13.8 kg/day

enter the minimum,


average, and maximum
lb. Average - 47.1 kg/day

quantities discharged


over the reporting
1c. Maximum - 161 kg/day

period, in the


corresponding spaces.


Page No. 18-17
-------
7 "'.CMTO'J'.iS PPO:EOL:RE: REPORTING OF SELF-MONITORING DATA
Paqe
No. 18-18

j



TRAINING
r' r ~ ¦** T ¦* * ' *" J7 7? -i -y • - — -
»
CTro
infcr:
'ATI0?;/C?ERv IN'-. COALS/SPECIFICATIONS
GUIDE NOTES
G. Suspended Solids,
2-
On the "Permit Condition"
2a.
Average suspended solids discharge over a

Final Effluent
1
|
line enter the average

30-consecutive-day period is specified as

(cont.)
}
and maximum quantities

80 kg/day (Table II)


j
permitted to be




1
I
discharged.
2b.
A maximum daily discharge limitation is not


i


specified in the permit (Table II). A dash


j
1


is therefore placed in the "maximum" space.


1
1
i

2c.
May already be entered by the Permit-issuing


i
i
t


authority.


3.
In the "No. Ex" column,
3a.
If none, enter a "0".



on the "Reported" line,





enter the number of
3b.
Since no maximum daily discharge limitation



times that the maximum

is indicated in the permit (Table II), a dash

;

daily discharge of

is placed in this space.

;

suspended solids (kg/day)



1

has been exceeded during



1
i

the reporting period.



3. "Concentration"
1.
On the "Reported" line
la.
Minimum - 12 mg/1

section.

enter the minimum,





average, and maximum
lb.
Average - 25.0 mg/1



suspended solids concen-





tration observed during
lc.
Maximum - 60 mg/1



the reporting period, in





the corresponding spaces.




2.
On the "Permit Condition"
2a.
Average suspended solids concentration over a



line enter the average

30-consecutive-day period is specified as



and maximum concen-

30 mg/1 (Table II).



trations specified in the





permit, in the appropriate





spaces.



-------
EFFLUENT "ONITORINS PROCEDURE: REPORTING OF SELF-MONITORING DATA
w : i_ T wl C. O „ i_, i 'w L
rii;r 'pi-
IMFC-RMATIOfi/CPERATIiJS COALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
G. Suspended Solids,
Final Effluent,
(cont.)
4. "Frequency of
Analysis"
column
5. "Sample Type"
column.
i 3. In the "No. Ex" column,
on the "Reported" line, ,
enter the number of times J
during the reporting j
period that the maximum
concentration allowed by
the permit has been
exceeded.
1.	On the "Reported" line
enter frequency of
analysis during the
reporting period.
2.	On the "Permit Condition"
line enter required
frequency of analysis, as
specified in permit.
On the "Reported" line
enter the type of sample
on which the analysis was
performed.
On the "Permit Condition"
line enter the type of
sample specified in the
permi t.
2b. Since no maximum concentration is specified in
the permit (Table II), enter a dash in this
space.
2c. May already be entered by the Permit-issuing
authority.
3a. If none, enter a "0".
3b. Since no maximum concentration is specified
in the permit (Table II), enter a dash in
this space.
la. Enter 2/7, indicating that the analysis was
performed twice weekly (Table I).
2a. Permit requires analysis twice weekly
(Table II). Enter 2/7.
2b. May already be entered by Permit-issuing
authority.
la. Enter "24-Hr. comp." since it is assumed in
this procedure that the data has been obtained
in accordance with permit requirements.
2a. Enter "24-Hr. comp." (Table II).
2b. May already be entered by the Permit-issuing
authority.
Page No. 18-19
-------
ppc-ceojie: reporting of self-monitoriijg data
Page No. 18-20
0" E"*.7!:;3 r-~CrE jLTES
STEP ?'ECUEr'CE
info?.:;atio;;/opcrating goals/specifications
TRAINING
GUIDE NOTES
G. Suspended Solids, j
Final Effluent. j
(cont.) I I
i

This portion of the Report Form, completed in accordance with assumed permit conditions, and the data of
Table I, is shown in Fig. 6 below:
CONCENTRATION
quantity
no.
161
25.0
47.1
2/7	24-Hr.CQmp
2/7 24-Hr.Comp
^ MG/L
1 KG/DAY
SUSPENDED SOLIDS
Fig. 6
Percent Removal
Suspended
Sol ids.
1. Computation.
2. "Quantity"
section
Calculate the percent
removal of suspended
solids for each pair of
influent and effluent
analyses made during the
reporting period.
On the "Reported" line
enter the minimum,
average, and maximum
percent removals in the
corresponding spaces.
la. Minimum - 66.61
lb. Average - 86.5/i
lc. Maximum - 93.0°:
II.H.l
(p-33 )
-------
Fr?:.U'!JT fiO?iITORi:^ ?*OCEDURE: REPORTING OF SELF-MONITORING DATA
D',CP.:,Tr.a ?";o::nu"^s
STEP SEQUENCE
I:; F 0 R - * A T10'; / 0 P E R A TI rG GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
H. Percent Removal
Suspended
Solids.(cont.)
3. "Concentration"
section.
t
' 2. On the "Permit Condition"
line enter the minimun
and average removals
required by the permit.
3. In the "No. Ex" column,
on the "Reported" line,
enter the number of times
that the minimum percent
removal required in the
permit was not obtained.
1. Enter dashes in the
"Units" space and in the
"No. Ex" space on the
"Reported" line.
2a. There is no minimum removal requirement in the
assumed permit conditions (Table II). Enter
a dash in this space.
2b. Average removal required over a 30-consecutive-
day period is 852 (Table II).
2c. May already be entered by the permit-issuing
authority.
3a. There is no minimum removal requirement in
the assumed permit conditions (Table II).
Enter a dash in this space.
3b. If none, enter a "0".
la. May already be entered by the Permit-issuing
authority.

This portion of the form, completed in accordance with assumed permit conditions, and the data of
T^ble I. is shown in Fig. 7 below:

Ul-JT
PARAMETER

{ 3 only)
IN'H.
quantity
'M-Sai

|'4 emrd only)
31*49
CONCENTRATION
M-!) *«-•<
FREQUENCY
e#.v.


MINIMUM
AVERAGE
MAXIMUM
' NO.
UNITS j Ex MINIMUM
AVERAGE MAXIMUM
NO °F
UNITS gx ANALYSIS
1
type 1

PERCENT REMOVAL
SUSPENDED SOLIDS
nCPORTSD
66.6
86.5
93.0
X




PENMi T
CONDITION
LO
CO
1

******

*—

Fi g. 7
Paae No. lft-21
-------
cFrLl!t::. MC:JITCr!"^ PROCEDURE: REPORTING OF SELF-MONITORING DATA
0PE-."T'PRCC"DL'.'*^S

STEP SEQUENCE

•'ATION/PPERATIHS GOALS/SPECIFICATIONS
TRAINING
GUIDE NOTES
I. Fecal Coliform.
(cont.)
4.
In the "No. Ex" column,
on the "Reported" line,
enter the number of times
during the reporting
period that the maximum
count allowed in the
permit has been exceeded.
4a.
4b.
If none, enter a "0".
Since no maximum count is specified in the
permit (Table II) enter a dash in this space.

4. "Frequency of
Analysis"
column
1.
Enter frequency of
analysis during the
reporting period on the
"Reported" line.
la.
Enter 2/7, indicating that the analysis was
performed twice weekly (Table I).


2.
Enter required frequency
of analysis, as specified
in permit, on the
"Permit Condition" line.
2a.
Permit requires analysis twice weekly
(Table II). Enter 2/7.

5. "Sample Type"
column.
1.
Ente*- type of sample on
which the analysis was
performed.
la.
Enter "Grab", since it is assumed in this
procedure that the sample has been obtained
in accordance with permit requirements.

This portion of the form, completed in accordance with assumed permit conditions, and the data of
Table I, is shown in Fig. 8 below:
H-«|i	

MRAMETCR

MeQUANTITY
 ' 1 NO.
MINIMUM J AVERAGE ] MAXIMUM . UNtTS
FECAL COLIFORM
RCPONTtD
******
******
******


110
220 j 540
j
N/100ML |j|
2/7 ! Grab
kimt
CONDI TION
******
******
******
"
_
¦
	
200 !
2/7 : grab
Fig. S
Page No. 18-23
-------
REPORTING OF SELF-MONITORING DATA
Page No. 18-24
J. Signature
MI KG GOALS/SPECIFICATIONS
I TRAINING
GUIDE NOTES
The completed form must
be signed by the ranking
elected official of the
municipality, or other
duly authorized
municipal employee.
Complete the four spaces
provided at the bottom
of the form.
Forward completed form
to permit-issuing
authority in accordance
with reporting
instructions specified
in permit.
2a. Name, title and signature of ranking elected
official or duly authorized employee, and date
of completion.
3a. The entire form, completed in accordance with
the permit conditions assumed for the purpose
of this procedure, is shown in Fig. 9.
-------
NATIONAL POLLUTANT DISCHARGE ELIMINATION SYSTEM
DISCHARGE MONITORING REPORT
Form Afpto rod
OMB NO. m*RD07$
City of Noname, Dept. of Environmental Services
184 Any Street
Noname, Anystate, 12345
lANj
Nfll ?34fifi7
PERMIT NUMBER
DHL
4952
REPORTING PERlOO FROM
DIS |
120-211 1 2 2- 2S
SIC
H 2«-Jft)
714 019
on
YEAR MO
OAY
47°20'4"
98°26' 11"
LAT
TUBE
LONGITUDE |

'IS-111 128
2*>


TO
714: 019
310


YEAR| MO
DAY

INSTRUCTIONS
1.	Provide dates foi period covered by this report in apace* atikcd "REPORTING PERIOD"
2.	Eater reported aiaiasa, average and auiaai valoea under "QUAMT1TT" nd"OOKCINTRATIOir'
ilk lha unite apecified for each piraiitu ^prapriate- Do not eater valike* ia bosea containing
asterisks. "AVERAGE" it average coopgted over actual tine discharge is operating. 'Hathi«"
and "WNDCUH" arc extreme value a observed duriag the reporting period
3.	Specify the number of analysed samples that esceed the ataxia am (and/or mirumtm a* tppropriata)
permit conditions in the columns labeled "Jfo. Ex." If none, enter "O".
4.	Specify frequency of analysis for esch parameter as Tie. analyses/No. day a. {•.%., "J/7" is equiva-
lent to 3 anafyaee partomod every 7 dayi.) If continuous enter "OOKT."
5.	Specify eaaaple type ("grab" or "— hr. conpoatfe") as ^pliceble. If frequency was continuous,
eater "HA".
6.	Appropriate si (nature i« required on bottom of this foia.
7.	Remove carbon and retain copy for your records.
8.	Fold along dotted lines, alaple and at ail Original to office apecified in permit
<«•»!>	-SS'TOI
PARAMETER
( 3 card only)
(4 card only)
M-481
CONCENTRATION
FREQUENCY
OF
ANALYSIS
sample
TYPg
RSPORTCO
FLOW
0.15
0.49
0.81
PCRMl T
CONDITION
MGD
Cont.
m.
RtPOR TSO
PH
5.4
9.2
PKKMl T
CONDITION
STANDARD
U.'IITS
6.0
9.0
Cont.
7/7 Grab
2/7 Grab
REPORT CO
BOD
11.0
37.5
94
PERMIT
CON TION
KG/DrtY
15
21
35
MG/L
2/7 24-Hr.C<«m
70
30
PERCENT REMOVAL
BOD
	S	
REPORTED
77
87.9
91
PERM! r
CONDITION
85
REPORTED
SUSPENDED SOLIDS
13.8
47.1
161
PERMIT
CONDITION
80
2/7 j24-Hr.C(ctd
KG/DAY
12
25.0
60
MG/L
30
2/7 i 24-Hr. Ctuno
2/7 124-Hr.C(mp
REPORTED
PERCENT REMOVAL
SUSPENDED SOLIDS
66.6
86.5
93.0
PIMMI T
CONDITION
85
REPORTED
FECAL COLI FORM
PERMI T
CONOJ Tl ON
REPORTED
110
220
540
200
N/100ML
ILL
ILL
Grab
CONDITION
NAME OF PRINCIPAL EXECUTIVE OFFICER
TITLE OF TNE OFFICER
OATE
!
j Doe
John
J. !
Mayor
7| 4
0| 9
3)0
[ LAST
FIRST
»• i
TITLE
YEAR
MO
DAY
/ certify that / am tmniti*r m&i the infounaUott contained in (ftrs
raport and that to A« beat ot my kaowtadga and beJiaf much Infor-
mation ia line, comp/efa, and accurate.
GRAB

5I5NATURE OP PRINCIPAL EXECUTIVE
OFFICER OR AUTHORIZED AGENT
Fiq. 9
1 OF 1
Paqe No. 18-25
-------
EFFLUENT MONITORING PROCEDURE: REPORTING OF SELF-MONITORING DATA
INTRODUCTION	Section I
TRAINING GUIDE NOTE
REFERENCES/RESCJRCES
A
Holders of discharge permits issued by the U. S.
Environmental Protection Agency are to report
self-monitoring data either on EPA Form T-40
(Fig. 10), or on EPA Form 3320-1 (Fig. 11).
Form T-40 will be used temporarily, and will be
furnished by EPA to municipalities for reporting
purposes. The T-40 form shown in Fig. 10 consists
of the 3320-1 form, which has been preprinted
for the reporting of data for basic parameters
common to all municipal wastewater discharges.
As information for each municipality is incor-
porated into EPA's computer system, form 3320-1
will replace Form T-40. For data reporting, a
municipality will then receive from EPA Form
3320-1 on which the effluent parameters specific
to that municipality will be computer preprinted.
Until that time, however, data for any additional
parameters to be reported which are not now in-
cluded in the preprint on T-40 will be entered
by the municipality, using as many additional
blank copies of the form as are required.
Completion of form T-40, is illustrated in this
procedure for the basic parameters, assuming that
permit conditions are as indicated in Table II.
Reporting of additional parameters would be done
in a manner similar to that illustrated.

Paqe No. 18-27
-------
Page No. 18-28
NATIONAL POLLUTANT DISCHARGE ELIMINATION SYSTEM
DISCHARGE MONITORING REPORT
Fom Appro red
OMBNO. 138-R007i
PERMIT NUMBER


LATITUDE
LONGITUDE
<»'»i) (afria <»»*»
REPORTING PERIOD FROM
n-iTi ias-»i no«»n
INSTRUCTIONS
1.	Provide dates for period covered by this report is spaces marked "REPORTING PERIOD"-
2.	Eater reported minimum. average and nuinia values under "QUANTITY" and "CONCENTRATION"
ift be units ^edfied for eadk paraeter as appropriate. Do not sater value* in boxes containing
asterisks. "AVERAGE" i» average computed over actual time discharfe is operating. "MAXIMUM"
and "MINIMUM" are extreme values observed daring the reporting period
1 Specify tike nimiber of analysed samples that exceed the maxim on (nd/ot minimum a* appropriate)
permit conditions in the columns labeled "No. Ex." If none, enter "O".
4.	Specify frequency of analyse for each parameter as No. snalyses/No. days, (e.g., "3/7" i* equiva-
lent Co 3 ana/yeee performed every 7 dmya.) If continuous enter "GONT."
5.	Specify sample type ( "grmb" or "	Sr. composite",) as ^plicable. If frequency was continuous,
sater "NA".
6.	Appropriate lipiiture ia required on bottom of this form.
7.	Remove carbon and retain copy for your records.
ft. Fold along dotted lines, staple and mail Original to office ^ecified in permit
		
PARAMETER
QUANTITY
tTAN0ARD
units
CONDITION
PCAMIT
CONDITION
KG/DAY
MG/L
PERMIT
CONOITtO*
PERMIT
CONDITION
KG/DAY
PERMIT
eOMO«TlO«
RMSMTID
PCRMIT
CONDITION
CONDITION
NAMg OF PRINCIPAL EXECUTIVE OFFICER
TITLE OF TNE OFFICER
*
HG/l
N/100MI
GRAB
/ certify Aaf / aei familiar with the infototetion contained in tfiie
raperf and that to ffie 6*et of my knowledge am/ Mid/ eucfr intor <
nation ie true, eomp/efe, and accurate.
TEAR MO
J.
SIGN A TU RE OF PRINCIPAL EXECUTIVC
OFflCER OR AUTHORIZED AGENT
T.JM4-N9
•10.
10
-------
NATIONAL POLLUTANT OlSC MARGE ELIMINATION SYSTEM
DISCHARGE MONITORING REPORT	Form Approved
OMBNO. IS6-R0073
J
PERMIT NUMBER


LATITUDE
LONGITUDE
(»~*>> taa-2»
I2S-27) m-f) 110-11)
REPORTING PERIOD. FROM
1
1
1
TO
1
1
1

YEAR
MO
OAY

YEAR
MO
DAY
INSTRUCTIONS
1.	Provide dates for period covered by this report in spaces marked "REPORTING PERIOD"-
2.	Eater reported minimum, average and maximum values under "QUANTITY" and "CONCENTRATION"
in the units specified for each parameter as ^propriate. Do not eater values in boxes containing
asterisk* "AVERAGE" is average computed over actual time discharge is operating. "MAXIMUM"
and "MINIMUM" are extreme values observed during the reporting pariod.
3.	Specify the number of analyzed samples that exceed the maximum (end/or minimtMn as appropriate)
permit conditions in the columns labeled "No. Ex." If none, enter "O".
4.	Specify frequency of analysis for each parameter as No. analyses/No. days, (e.g., "3/7" it equiva-
lent to 3 analyses performed every 7 days.) If continuous enter "CONT."
5- Specify sample type ("grab" or - hr. composite") as applicable. If frequency was continuous,
enter "NA".
6.	Appropriate signature is required on bottom of this form.
7.	Remove carbon and retain copy for your records.
8- Fold along dotted lines, staple and mail Original to office specified in permit.

PARAMETER

(3 cmr4 only)
(SS-«S>
QUANTITY
I4S*SS> 1S*-»II



CONCENTRATION
- 46'99l
(62*4)1
FREQUENCY
SAMPLE


MINIMUM
AVERAGE
MAXIMUM
UNITS
NO.
EX
MINIMUM
AVERAGE
MAXIMUM
UNITS
NO.
EX
OF
ANALYSIS
TYPE


REPORTED









J



PERMI T
CONDITION




¦




u



REPORTED









1 i
i t


PERMIT
CONOI T10N




I




]A



REPORTED









J



PERMIT
CONDITION




¦




m



REPORTED










r-
1
i


PERM! T
CONDITION




£



m




REPORTED













PERMI T
CONOlTION




II




m



REPORTED









H



PERMIT
CONOI TION




i








REPORTED














PERMIT
CONDITION




ft








REPORTED




i








PERM! T
CONOlTION




2]





NAME OF PRINCIPAL EXECUTIVE OFFICER
TITLE OF THE OFFICER
|
DATE



i





|









| t
1
|
matton it true, complete, and accurate.
| SIGNATURE OF PRINCIPAL EXECUTIVE
LAST FIRST
Ml
TITLE
j YEAR

MO
OAY


OFFICER OR AUTHORIZED AGENT
EPA Fern 3320-1 (*0-72)
Fig. 11
Page No. 18-29
-------
EFFLUENT MONITORING PROCEDURE: REPORTING OF SELF-MONITORING DATA
EDUCATIONAL CONCEPTS - MATHEMATICS
Section II
TRAINING GUIDE NOTE
REFERENCES/RESOURCES
NOTE: In all of the calculations in this section,
rules for computation as given by Crumpler
and Yoe were followed. Crumpler, T. B. and
Yoe, J. H. Chemical Computations and Errors.
Wiley and Sons. N.Y. 1940.
Cnmputation of Quantities of BOD discharged in Final
Effluent.
Pertinent reported data from Table I is listed in
the first three columns of the Table below. In
column 4 the flow has been converted from gpd
to MGD. The quantity of BODj- is obtained by multi-
plying the value in column 3 (mg/1) by the value
in column 4 (MGD), and then multiplying the result
by the factor 3.78.
This is expressed in mathematical form as
Kg/day = MGD x mg/1 x 3.78
Example:
On September 3, Kg/day = 0.33 x 16 x 3.78 = 20
Date
Flow
gpd
BODc
mg/1
; Flow
1 MGD
BODr
Kg/day
3
326,900
16
0.33
20
6
458,500
15
0.46
26
9
146,000
20
0.146
11.0
12
519,200
20
0.52
39
16
708,900
35
0.71
94
20
272,900
19
0.27
19
23
761,800
20
0.76
57
26
291,600
20
0.29
22
29
525,600
25
0.53
50

Total
190

338

Average B0D5 =
190
9
21 mg/1


Average BOD^ =
333
9
37.5 Kg/day

Crumpler, T. B. and Yoe,
J. H. Chemical Computations
and Errors. Wiley and Sons.
N.Y. 1940.
Page No. 18-30
-------
EFFLUENT MONITORING PROCEDURE: REPORTING OF SELF-MONITORING DATA
EDUCATIONAL CONCEPTS - MATHEMATICS
Section II
TRAINING GUIDE MOTE
REFERENCES/RESOURCES
F.I
Computation of percent BOD^ removals.
Pertinent reported data from Table I is listed in
the first three columns of the Table below. The
percent BOD,- removal for each day appears in
column 4.
The percent removal is obtained by subtracting the
concentration of BOD5 in the final efficient from
that in the plant influent, dividing this
difference by the concentration of BOD5 in the
influent, and multiplying the result by 100. This
can be expressed in mathematical form as follows:
*bod5	. mnuent	BOD, N/1) , 100
Example:
On September 3, % BOD Removal = 170j7016 x 100 = 91%
BODs-mg/l
Date	Inf.	Eff. % Removal
3	170	16	91
6	160	15	91
9	200	20	90
12	190	20	89
16	150	35	77
20	170	19	89
23	150	20	87
26	190	20	89
29	_190	_2£	87
Total	1,570	190
Average	174	21
Average	BOD5 Removal -	x 100 = 87.9%
Page No. 18-31
-------
EFFLUENT MOMITORING PROCEDURE: REPORTING OF SELF-MONITORING DATA
EDUCATIONAL CONCEPTS - MATHEMATICS	Section II
TRAINING GUIDE MOTE
REFERENCES/RESOURCES
.1
Computation of Quantities of Suspended Solids
Discharged in Final Effluent.
Pertinent reported data from Table I is listed in
the first three columns of the Table below. In
column 4 the flow has been converted from gpd to
MGD. The quantity of suspended solids is obtained
by multiplying the value in column 3 (mg/1) by the
value in column 4 (MGD), and then multiplying the
result by the factor 3.73. This is expressed in
mathematical form as:
KG/day = MGD x mg/1 x 3.78

Example: On September 3, Kg/day = 0.33 x 12 x 3.78 = 15
Date
Flow
T
.S.S.
Flow
T.S.S.

gpd

mg/1
MGD
Kg/day
3
326,900

12
0.33
15
6
458,500

16
0.46
23
9
146,000

25
0.146
13.8
12
519,200

25
0.52
49
16
708,900

60
0.71
161
20
272,900

19
0.27
19
23
761,800

23
0.76
66
26
291,600

20
0.29
22
29
525,600

25
0.53
50
Total

225

424
Average T.S.S. =
225
9
= 25.0 mg/1

Average T.S.S. =
424
9
1)
¦C*
kg/day

Page No. 18-32
-------
EFFLUENT MONITORING PROCEDURE: REPORTING OF SELF-MONITORING DATA
EDUCATIONAL CONCEPTS - MATHEMATICS
Section II
TRAINING GUIDE NOTE
REFERENCES/RESOURCFS
Computation of percent suspended solids removal.
Pertinent reported data from Table I is listed in
the first three columns of the Table below. The
percent suspended solids removal for each day
appears in column 4.
The percent removal is obtained by subtracting the
concentration of suspended solids in the final
effluent from that in the plant influent, dividing
this difference by the concentration of suspended
solids in the plant influent, and multiplying the
result by 100. This can be expressed in mathe-
matical form as follows:
T r r „ Influent T.S.S. (mq/1) - effluent T.S.S. (mg/1) lnn
Removal = 	Influent T.S.S. (mg/1)	 	 x 100
Example:



On September 3, !
I T.S.S.
17
removal = —
Date
T.S.S
Inf.
.-mg/1
Eff.
% Removal
3
171
12
93.0
6
168
16
90.5
9
200
25
87.5
12
198
25
87.4
16
180
60
66.6
20
170
19
88.8
23
186
23
87.6
26
195
20
89.7
29
195
25
37.2
Total
1,663
225

Average
185
25

171
x 100 = 93.0°;
185-25
Average % Removal = —x 100 = 86.5%
Page No. 18-33
-------
EFFLUENT MONITORING PROCEDURE: REPORTING OF SELF-MONITORING DATA
EDUCATIONAL CONCEPTS - MATHEMATICS
Section II
TRAIN I KG GUIDE NOTE
REFERENCES/RESOURCES
1.1
Computation of the Geometric Mean
Pertinent reported data from Table I is listed in
the first two columns of the Table below. The
logarithm of the reported Coliform value appears in
column 3. Note that two-place logarithms are used
in this calculation. That is, the mantissa of the
logarithm (the numbers to the right of the decimal
point) contains only two numbers. Two-place
logarithms are adequate since the coliform values
are reported only to two significant figures.
The logarithms in column 3 are added, the total is
divided by the number of values reported, and the
anti-logarithm of the quotient is obtained. This
is the geometric mean. It is reported to two
significant figures.
Date	Fecal Coliform Log of Fecal
N/100 ml
Coli form
3
6
9
12
16
20
23
26
29
350
540
180
170
220
240
110
130
280
2.54
2.73
2.26
2.23
2.34
2.38
2.04
2.11
2.45
Total
21.08
The antilogarithm of 2.34
is 220. This is the
geometric mean. If the
antilogarithm did not end
with a zero, the number
would he rounded to the
nearest ten for reporting
purposes.
Page No. 18-34
-------
EFFLUENT MONITORING PROCEDURE: REPORTING OF SELF-MONITORING DATA
RECORDS AND REPORTS	Section IX
TRAINIMG GUIDE NOTE
REFERENCES/RESOURCES
C.T.I.
Reporting of minimum and/or maximum values may or
may not be required by the permit-issuing author-
ity. If not required, a dash or an asterisk may
already be entered in either or both of these
spaces. The same is true for all of the other
parameters shown, with the exception of pH, for
which minimum and maximum values must be reported.

C.1.3.3a
Preprinted forms may already have either a dash
or an asterisk in this space. This also applies
to all other cases in this procedure where the
entry of a dash in a space is specified.

Page No. 18-35
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