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Course Guide Che 3131l

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Course Guide Che 3131l

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crystahsoo

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COURSE GUIDE IN

ENVIRONMENTAL SCIENCE AND ENGINEERING FOR

CHE(LAB)

CHE 3131L

CHEMICAL ENGINEERING DEPARTMENT

SCHOOL OF ENGINEERING AND ARCHITECTURE

Contents Page

Title Page 1

Course Overview 3
Course Study Guide and House Rules 5

Study Schedule 7

Assessment and Evaluation Guide

General Requirements 9
Formative Assessment Guide 9
Evaluative Assessment Guide 10

Technological Tools 10
Grading System 11

Course References 11

Facilitator Contact Details 12

COURSE LEARNING OUTCOMES

The course CHE 3131L: Environmental
Science and Engineering for CHE
Laboratory prepares you, as a future
chemical engineer, to demonstrate all
the intended learning outcomes. At the
end of the course you should be able
to:

CLO 1: Exhibit an in-depth

understanding on the proper
procedures of water sampling and the
different standard methods of
environmental laboratory tests.

CLO 2: Demonstrate the different

sampling methods, preservation and
ENVIRONMENTAL storage of both water and wastewater
SCIENCE & ENGINEERNG samples.

(LABORATORY) CLO 3: Perform standard procedures in

determining water quality parameters
as well as procedures for other related
environmental laboratory tests.

CLO 4: Analyze the water quality

parameters and characterize the water
samples from community sources.

In this course, the would-be Chemical Engineers learn the proper procedures used in the
water sampling and the different standard methods of environmental laboratory tests for
water and wastewater employed in monitoring the physical, chemical and biological quality
of water and wastewater.

Outcomes-based education dictates that only when you can demonstrate the course
learning outcomes by the end of this course, can you be given a passing mark. *Laboratory
work refers to the report on the analysis of sample data and DIY experiments that can be
performed using available household chemicals and resources.

The modules that form the building blocks to help you attain the course learning outcomes
are as follows:

MODULE 1: Standard laboratory procedures in sampling and preparation

MODULE 2: Water characterization

MODULE 3: Standard methods for the analysis of water and wastewater samples

The key to successfully finish this online course relies heavily on your self-discipline and time
management skills. This module was prepared for you to learn diligently, intelligently, and
independently. Keeping yourself motivated to follow the schedules specified in the learning
plan, maintaining excellence in the expected student outputs, and mastering the different
technologies and procedures required in the delivery and feedback for this course, will instill
in you important qualities you will need in the future as an engineer practicing your
profession. The following course guides and house rules are designed for you to practice
decorum consistent with standards expected within a formal academic environment. These
guides shall lay the groundwork for consistency, coherence, cooperation, and clear
communication among learners and instructors throughout the conduct of this course:
1. MANAGE YOUR MINUTES. Create a study routine and stick to it. Keep requirement
deadlines and study schedules always in mind by providing visual cues posted in your
place of study or listed in your reminders (electronically, online, or on paper).
Remember that there are other daily activities that take up your time, not to mention
other courses you may be concurrently taking. Choose a time of day when you are
most likely to maximize learning. Communicate your schedule to other members of
your household so they could help you keep it. It would also help to prepare a
dedicated space in your residence conducive for learning.

2. MIND YOUR MANNERS. Treat the distance learning environment as an academic

space not too different from a physical classroom. Do not do in the distance learning
environment, acts you would not normally do in a face-to-face classroom set up.
Avoid asking questions that have already been answered in the lessons or in the
instructions previously discussed or provided. This reflects your poor focus and
uninspired preparation for this course. Practice Conscientious Habitual Etiquette in
group chats, open forums, and similar electronic venues.
a. Use appropriate language and tone, correct grammar and spelling, and
complete sentences acceptable in an academic forum. Avoid text-speak,
slang, and all caps in your posts.
b. Express your opinions politely and do not dominate the conversation.
c. Avoid lengthy as well as offensive posts by sticking to the topic of the
discussion.
d. Take time to understand the salient points of the discussion, and provide
meaningful and well-thought responses to the posts of other participants.
e. For a live meeting or video/voice conferencing set-up, mute your microphone
when you are not speaking to keep the focus on the main speaker.

3. MASTER THE MEDIUM. The distance learning courses will be delivered making use of
the institutional Google Suite account of Saint Louis University. It would be worthwhile
on your part to devote some time and effort to learn the applications you will need
to access your course materials, interact with me and your classmates, and submit
course requirements. Applications of note are Google Classroom, Google Drive, and
Google Meet. There are also available alternatives to Microsoft Office tools you might
want to explore. Certain requirements will require you to take a video on your smart
phone, save it, and submit it electronically. Work on this skill as well. If you are offline,

4. MAKE MASTERPIECES. Go beyond minimum requirements. The course learning

outcomes will serve as a guide to the minimum expected competencies you are to
acquire at the end of this course. It does not limit you from performing beyond it.
Keep in mind that the quality of your work reflects the amount of thought and care
you put into the process of completing it. It provides a very tangible measure of how
much of the competencies you have developed and fully obtained throughout this
course.

5. CONNECT CONSTANTLY. There are more than sufficient online and offline modes to
ensure that you are well informed and provided on time with the needed learning
materials, instructions, requirements, and feedback either from me or from your
classmates. Exhaust all means possible to keep in touch and updated. My contact
details can be found at the latter part of this document and will be made available
and widely disseminated to enrolees of this course.

6. OBSERVE ORIGINALITY. Your course outputs will largely be submitted in electronic

form. It is going to have a highly traceable and comparable digital footprint that can
be easily checked for originality. Cite your sources properly for referenced statements
you decide to use in your own work. Attribute statements by persons other than you
by using terms like according to, he said/she said, and the like.

7. INSTIGATE INDEPENDENCE. You are the focus of this course. Nobody else. All
assessment and evaluation tools in this course are designed to measure your
competence and not anybody else’s. You may use all resources at your disposal,
and ask other people for advice. In the end however, it is going to be your
independent work that will be judged against the standards set for this course. The
only way for you to maximize this course to your advantage is to learn as much from
it as an individual. Make it count.

Additional Guidelines for Offline Students

If you are a student opting for the offline mode of distance learning, you will be tasked to
send back the accomplished requirements at given stages of the course through express
mail correspondence on or before the scheduled date to me. Make sure you will follow it up
with me through text or any other media available for you.

While waiting for my feedback of your accomplished requirements, continue doing the task
in the succeeding units of the module. If needed, do not hesitate to keep in touch with me
through any available means. Remember, if there is a will, there is a way.

Below is the complete weekly schedule for the attainment of the topic learning outcomes
vis-a-vis the activities. This contains also the schedule of the deadlines of the submission of
the accomplished course requirements or assignments and the examination.

Dates TOPIC LEARNING OUTCOMES ACTIVITIES

MODULE 1 Standard laboratory procedures in sampling and preparation
UNIT 1 Standard laboratory guidelines and procedures
Week 1 TLO 1: Understand the standard Introduction to the course
laboratory guidelines and Discussion on the significance of the
procedures course outline
UNIT 2 Water sampling, storage and preservation
Week 2 TLO 2: Familiarize the standard Discussion of Theory
operating procedures in water Video Demonstration of procedures
sampling, storage and preservation.

MODULE 2 Water characterization

UNIT 1 Physical and chemical parameters of water
Week 3 Discussion of related regulations:
 DAO 2016-08. Water Quality
TLO 3: Measure the different physical Guidelines and General Effluent
and chemical parameters of water Standards of 2016.
such as pH, temperature, turbidity  DOH Administrative Order 2017 –
and conductivity. 0010. Philippine National
Standards for Drinking Water of
2017
UNIT 2 Solids Analysis
Week 4 Discussion on the procedure for
and 5 TLO 4: Perform the standard determination of solids content of
operating procedures in determining water
the amount of solids present in Video Demonstration of procedures
water. Design of experiments for DIY testing

Module 1 & 2 Evaluative Assessment

Week 6
*Prelim Examination
MODULE 3 Standard methods for the analysis of water and wastewater samples
UNIT 1 Argentometric Method
Week 7 Discussion of Theory
TLO 5: Demonstrate the procedure
Video Demonstration of procedures
in determining the chloride and iron
Sample calculations
content of water.
Analysis of sample data
UNIT 2 Titrimetric Method
Week 8 TLO 5: Demonstrate the procedure in Discussion of Theory
determining the chloride and iron Video Demonstration of procedures
content of water. Sample calculations

TLO 7: Measure the amount of

Ammonia and Nitrates level in water.
UNIT 3 UV-VIS Spectrophotometric Screening Method
Week 9 TLO 5: Demonstrate the procedure in
determining the chloride and iron
content of water.
Discussion of Theory
TLO 6: Demonstrate the procedures
in determining the hardness of water. Video Demonstration of procedures

TLO 7: Measure the amount of Sample calculations

Ammonia and Nitrates level in water.
Analysis of sample data
TLO 8: Perform the standard
procedures in determining Sulfates
and Phosphates content of water.
UNIT 4 Turbidimetric Method
Week 10 Discussion of Theory
TLO 8: Perform the standard Video Demonstration of procedures
procedures in determining Sulfates Sample calculations
and Phosphates content of water. Analysis of sample data

UNIT 5 Ascorbic Acid Method

Week 11 Discussion of Theory
TLO 8: Perform the standard Video Demonstration of procedures
procedures in determining Sulfates Sample calculations
and Phosphates content of water. Analysis of sample data

Module 3: Units 1-5 Evaluative Assessment

Week 12
*Midterm Examination
UNIT 6 Azide Modification Method and 5-Day BOD Test
Week 13 TLO 9: Demonstrate the procedures Discussion of Theory
in determining the amount of Video Demonstration of procedures
Dissolved Oxygen (DO) present in Sample calculations
water Analysis of sample data
TLO 10: Execute the standard
procedures in determining the
Biochemical Oxygen Demand of
water.
UNIT 7 Open Reflux Method
Week 14 Discussion of Theory
TLO 11: Demonstrate the methods
Video Demonstration of procedures
in determining the Chemical
Sample calculations

UNIT 8 Multiple Tube Fermentation Technique

Week 16 TLO 12: Measure the total coliform Discussion of Theory
and fecal coliform present in Video Demonstration of procedures
water. Sample calculations
Analysis of sample data
Compilation of reports from the analysis of sample laboratory data.
Week 17
Comprehensive Final Exam
a. Multiple choice
Week 18 b. Problems solving
c. Submit video recording of DIY experiments

ASSESSMENT AND EVALUATION GUIDE

The course modules rely on formative and summative assessments to determine the progress
of your learning in each module. To obtain a passing grade in this course, you must:

1. Read all course materials and answer the pre-assessment quizzes, self-assessment
activities, and/or reflection questions.
2. Participate in online discussion forums.
3. Submit all assignments and graded quizzes.
4. Take the Midterm and Final Examination

Formative Assessment Activities

Formative assessments for this course are applied to ungraded activities that are used to
monitor your learning experience and provide feedback to improve both your learning
approach as well as my instructional approach.
• You are required to answer the pre-assessment quizzes, self-assessment activities,
and reflection questions but your scores in these activities will not be included in the
computation of your final grade.
• The reflection questions are designed to help you to critically analyse the course
readings for better understanding while the pre-assessment quizzes and self-
assessment activities are designed as a review management tool to prepare you
for the graded quizzes and examinations.
• Successfully answering formative activity questions and requirements will serve as
prompts to tell you if you need to study further or if you may already move forward
to the next unit of the module.
• The completeness of your answers to the pre-assessment quizzes, self-assessment
activities, and reflection questions will still be checked and will still be part of your
course completion. Hence, these must be answered.

Evaluative Assessment Activities

The evaluative assessments are graded activities designed to determine if your acquisition
of learning and performance in tests is at par with standards set at certain milestones in this
course.

A. Quizzes, Examinations, and Assignments

Graded quizzes, examinations, and assignments are essential to determine whether your
performance as a student is at par with standards/goals that need to be achieved in this
course. The scores obtained from each of the graded activities will contribute to your final
grade, the weights of which are presented in the grading system described in the
succeeding sections of this text. Direct scoring can be used on straightforward requirements
like short answers and multiple choice responses, while scoring rubrics will be provided for
answers that are typically lengthy and involve a more complex level of thinking on your part.

B. Final Course Requirement

You will need to accomplish a compilation of reports and comprehensive exam as a final
requirement for the course. A presentation explaining the experiments designed at home
will translate learned concepts and principles into the expected outcomes for this course.
For online students, a live presentation will be scheduled on Google Meet. For offline
students a recorded and saved presentation will be accommodated for submission on a USB
flash drive. A separate rubric will be used for the presentation and will constitute 20% of the
total score of the final examination.

Technological Tools

To be able to accomplish all the tasks in this course, you will need a computer or a laptop
with the following software applications: Word Processing, Presentation, and Publication for
requirements that do not require online access. A smart phone with video recording and
editing features will also be used for activities that will require you to record videos for saving
and submission.

If you are a student offline, the delivery of instructions and requirements will be primarily
through express mail correspondence of printed modules and saved digital content on a
USB flash drive. Feedback and clarifications will be facilitated through text messaging and
voice calls; hence, you need to have regular access to a cell phone. If you need to call, or

Grading System
Activity Weight
PRELIM GRADE (PG)
CS (online discussion, quiz) 10%
Laboratory reports 20%
Examination 60%

MIDTERM GRADE (MG)

CS (online discussion, quiz) 10%
Laboratory reports 20%
Examination 60%

TENTATIVE FINAL GRADE (TFG)

CS (online discussion, quiz) 5%
Laboratory reports 15%
Examination 70%

Final Grade: (PG + MG + TFG)/ 3 = 100%

COURSE REFERENCES
A. Main Reference/s
1. APHA (2017) Standard Methods for the Examination of Water and
Wastewater 23rd Edition. American Public Health Association; American
Water Works Association; Water Environment Federation.

B. Books
1. Davis, Mackenzie (2012) Introduction to Environmental Engineering, 5th Ed.
McGraw-Hill Education
2. Enger, Eldon D. (2013) Environmental Science: A Study of Interrelationships,
13th Ed. McGraw-Hill Education
3. Kiely, Gerard (1997). Environmental Engineering. Michigan: McGraw-Hill
4. Metcalf & Eddy Inc. (1991). Wastewater Engineering: Treatment, Disposal and
Reuse, 3rd Edition. McGraw-Hill Education
5. Nazaroff (2001). W.W. Environmental Engineering Science. Michigan: Wiley
6. Vesilind, P.A. (2010). Introduction to Environmental Engineering, 3rd Edition.
Cengage Learning
7. Sawyer, Clair (2003). Chemistry for Environmental Engineering and Science,
5th Ed. McGraw-Hill Education

CONTACT INFORMATION
Engr. Melissa May M. Boado
CHE 3131L Course Facilitator

Cellphone : +63927 124 7124

SLU local extension number : Chemical Engineeing, loc. 391
Institutional email address : [email protected]

Revised and updated: 10 August 2020

Introduction

Standard Operating Procedures (SOP) are instructions which will describe, in detail, how to
perform laboratory process or experiments safely and effectively in using
materials/apparatus whether a chemical, radioactive and biological, and physical hazards
that may occur.

UNIT 1 Standard laboratory guidelines and procedure

Laboratory Guidelines

1. The class will be divided into groups that will be collaborating together in the
laboratory and in writing the laboratory reports. Each group is required to submit only
one lab report. (Note: If other member/s have not cooperated during the experiment
or in the writing of laboratory report, name/s should not be included on the
manuscript.)
2. Always bring lab manual, calculator, lab notebook, and laptop (as preferred by the
student).
3. All laboratory data should be logged on a class notebook to be submitted by the end
of the period to the instructor. Laboratory Reports will be submitted on a scheduled
date and time as instructed by the instructor.
4. All reports should strictly follow the outline below:
 Title Page: It includes course title, experiment title, month and year the
experiment performed and names of group members. A format will be posted.
 Abstract: A brief statement covering the important objectives, background
materials, procedures, result, and conclusions. APA format should be applied.
 Introduction: A general introduction that will provide a short overview or the
theoretical background of the study.
 Design and Methodology: Shows an experimental design and specific
procedures or techniques used in the experiment.
 Results and Discussion: This section includes a presentation of data gathered
and calculated in tabular or graphical format. Proper labelling should be
observed.
 Conclusions and Recommendations: Interpretation of the findings or results
with recommendations that are specific and realistic should be discussed in
this section.

Property of and for the exclusive use of SLU. Reproduction, storing in a retrieval system, distributing, uploading or posting online, or transmitting in any form or by any
means, electronic, mechanical, photocopying, recording, or otherwise of any part of this document, without the prior written permission of SLU, is strictly prohibited. 13
Appendices: Statistical analysis, calculations, answers to questions are

presented in this section.
 References: Citations in the manuscript should be properly documented here.
APA Format should be applied.
Lab Safety and Etiquette
Before the class period/experiment period
 Before entering the laboratory, students should wear properly the lab gowns.
 Student should wear full sleeves clothes or lab gown and full-closed shoes.
 Wear safety glasses and gloves when recommended by the faculty before
conducting experiment.
 Bags and coats should be placed on designated areas away from the working areas.
 Listen to instructions before starting with the experiment.
During the class/experiment period
 Eating, drinking, playing, or application of cosmetics (including hand lotion, etc.) are
strictly prohibited.
 Check broken apparatus or glasswares before starting the experiment.
 Place broken glasswares in specially marked containers.
 Take an amount of chemicals from the reagent bottles that will only be needed in the
experiment.
 Never pour a chemical back into a reagent bottle.
 Properly wash your hands after contact with hazardous materials.
After the class/experiment period
 Return apparatus and chemicals to the laboratory custodian counter.
 Before leaving the working area, be mindful of your belongings, check for litters to be
disposed in marked containers.

Procedure for cleaning of glassware in laboratory

Glassware Cleaners
1. Clean the equipment thoroughly with soap and water for basic cleaning. You may
need to use a wire brush to remove some residue. Detergent using bottle brushes and
scouring pads can be used as needed.
2. After cleaning, rinse the glassware with running tap water. When test tubes,
graduates, flasks and similar containers are rinsed with tap water, allow the water to
run into and over them for a short time, then partly fill each piece with water.
3. Thoroughly shake and empty at least six times and ensure that all soap residue is
removed.
Note:
 Do not use cleaning brushes that are so worn that the spine hits the glass. Serious
scratches may result. Scratched glass is more prone to break during experiments. Any
mark in the uniform surface of glassware is a potential breaking point, especially when
the piece is heated. Do not allow acid to come into contact with a piece of glassware
before the detergent (or soap) is thoroughly removed. If this happens, a film of grease
may be formed.
 To prevent breakage when rinsing or washing pipets, cylinders or burets, be careful
not to let tips hit the sink or the water tap.
Sterilizing Contaminated Glassware
 Autoclave glassware or sterilize it in large steam ovens or similar apparatus. If viruses
or sporebearing bacteria are present, autoclaving is absolutely necessary.

UNIT 2 Water sampling, storage and preservation

Water quality standards are put in place to ensure the efficient use of water for a designated
purpose. Water quality analysis is to measure the required parameters of water, following
standard methods, to check whether they are in accordance with the standard.

Why is it required to do water quality analysis?

Water quality analysis is required mainly for monitoring purpose. Some importance of such
assessment includes:
1. To check whether the water quality is in compliance with the standards, and hence,
suitable or not for the designated use.
2. To monitor the efficiency of a system, working for water quality maintenance.
3. To check whether up gradation / change of an existing system is required and to
decide what changes should take place.
4. To monitor whether water quality is in compliance with rules and regulations.

Sampling of Water for Analysis:

A common cause of error in water quality analysis is improper sampling. The results of a water
quality analysis of a sample show only what is in the sample. For the results to be meaningful,
the sample must be representative i.e., it must contain essentially the same constituents as
the body of water from which it was taken.

The objective of sampling is to collect a portion of material small enough in volume to be

transported conveniently and yet large enough for analytical purposes while still accurately
representing the material being sampled.

The complexity of water quality as a subject is reflected in the many types of measurements
of water quality indicators. The most accurate measurements of water quality are made on-
site, because water exists in equilibrium with its surroundings. Measurements commonly
made on-site and in direct contact with the water source in question include temperature,'
pH, dissolved oxygen, electric conductivity, etc. More complex measurements are often
made in a laboratory requiring a water sample to be collected, preserved, transported, and
analyzed at another location.

Requirements for Sampling:

 Meet the requirements of the sampling program.
 Handle the sample carefully so that it does not deteriorate or become contaminated
or compromised before it is analyzed.
 Ensure sampling all equipment are clean and quality assured before use.
 Use sample containers that are clean and free of contaminants.

Sample Collection bottles, Size and Materials:

The methods that will be followed will determine the type of bottles used. For example,
samples for metals’ analyses are usually collected in plastic bottles, while analyses for volatile
organics and pesticides are collected in glass containers. Bottles used to collect samples for
bacteria should be sterilized. Certain analysis like volatile organics and radon require vials
that are to be filled leaving no head space, which keeps these analytes dissolved in the
water, preventing them from escaping into the air. Additionally, some analyses require
samples to be collected in amber colored bottles. These darker bottles are for analytes that
break-down in sunlight, which helps keep these contaminants from breaking down while in
transit to the laboratory for analysis. The size of the container is important to ensure enough
sample to run the analysis needed.

Water Sampling Techniques:

Water sampling can be done in any of the following three methods depending on test
requirements:
 Grab sampling
Grab Samples are samples collected at a particular time and space. They
represent the composition at that time and place. When a source is known to vary in
time e.g.in case of waste effluents, grab samples collected at suitable time intervals
and analyzed separately.

 Composite sampling
Composite samples are a mixture of grab samples collected at one sampling
point at different times. The composite samples are useful for observing values.
Individual samples are collected in wide mouth bottles every hour and mixed in
volume proportional to the flow or by using specially designed automatic sampling
devices.

Advantages of Composite Samples:

- reduced costs of analyzing a large number of samples.
- more representative samples of varied conditions.
- larger sample sizes when amounts of test samples are limited.

Disadvantages of Composite Samples

- loss of analytic relationships in individual samples.

 Integrated sampling
Integrated samples are a mixture of grab samples collected from different
points simultaneously and mixed in equal volumes.

Surface Water Sampling Techniques:

When the water source is accessible
 Rinse the sampling vessel with water on site 3-4 times.
 Care must be taken to avoid contaminating water to be sampled during rinsing.
 Submerge the sampling vessel gently, fill it with the water sample and close it tightly.
 If the collected water sample may be frozen, leave some space for expansion
equivalent to about 10% of the sampling vessel.

When the water source is inaccessible

A rope 'attached to the bucket are often used. Scoops with adjustable shafts are
convenient. Items made of synthetic resins such as polypropylene can also be used.

Ground Water (from well) Sampling Techniques

A bailer in is a hollow tube used to retrieve groundwater samples from monitoring wells.
Bailers are tied to a piece of rope or a piece of wire and lowered into the water column.
Once lowered, the bailer uses a simple ball check valve to seal at the bottom in order to pull
up a sample of the groundwater table.

Groundwater Sampling Bailers

Proper Labelling
Proper labelling prevents sample misidentification and ensures the responsibility and
accountability of the collector. The sample container should be labeled properly,
preferably by attaching an appropriately inscribed tag or label. Alternatively, the botthe
can be labeled directly with a water-proof marker. Barcode labels are also available
nowadays.

Information on the sample container or the tag should include at least:

i. Sample code number (identifying location)
ii. Date and time of sampling
iii. Source and type of sample
iv. Pre-treatment or preservation carried out on the sample
v. Any special notes for the analyst
vi. Sampler’s name

Sample Preservation:
There is usually a delay between the collection and analysis of a sample. The nature of the
sample can be changed during this period. Therefore proper preservation is required in the
way to laboratory after collection, and in the laboratory up to when analysis starts.

Complete and unequivocal preservation of samples, whether domestic wastewater,

industrial wastes, or natural waters, is practically impossible as because - complete stability
for every constituent never can be achieved.

At best, preservation techniques only retard chemical (especially, hydrolysis of

constituents) and biological changes that inevitably continue after sample collection. No
single method of preservation is entirely satisfactory; the preservative is chosen with due
regard to the determinations to be made.

Commonly used preservation methods are - pH control, chemical addition, the use of
amber and opaque bottles, refrigeration, filtration, and freezing.

Make an interactive video presentation to show on how you are to collect water sample
from the source where you usually get for your household activities. Observe the correct
way or standard procedure during sample collection up to preservation. And in your video,
include also a description on the details of your water source, from where it is coming from.

Water movement in our environment can be explained through the hydrological cycle. As
it moves from one state to another, it acquires impurities when it comes into contact with
materials in the air and on or beneath the earth’s surface. In addition, human activities
contribute further impurities in the form of industrial and domestic wastes, agricultural
chemicals, and other less obvious contaminants.

Standards for water quality are put in place to ensure the efficient use of water for a
designated purpose. The objective is to measure the required parameters of water, following
standard methods, to check whether they are in accordance with the standard.

Water quality analysis is required mainly for monitoring purposes. Some importance of such
assessment includes:
1. To check whether the water quality is in compliance with the standards, and hence,
suitable or not for the designated use.
2. To monitor the efficiency of a system, working for water quality maintenance.
3. To check whether upgradation/change of an existing system is required and to
decide what changes should take place.
4. To monitor whether water quality is in compliance with rules and regulations.

Experiment 1
Determination of temperature, pH, turbidity and conductivity

Principle:
pH value of water indicates the hydrogen ion concentration in water and concept of pH
was put forward by Sorenson (1909). pH is expressed as the logarithm of the reciprocal of
the hydrogen ion concentration in moles/liter at a given temperature. The pH scale
extends from 0 (very acidic) to 14 (very alkaline) with 7 corresponding to exact neutrality at
25°C. pH is used in the calculation of carbonate, bicarbonate and CO2, corrosion and
stability index etc. While the alkalinity or acidity measures the total resistance to the pH
change or buffering capacity, the pH gives the hydrogen ion activity.

pH can be measured colorimetrically or electrometrically. Colorimetric method is used only

for rough estimation. It can be done either by using universal indicator or by using pH
paper. The hydrogen electrode is the absolute standard for the measurement of pH. They
range from portable battery operated units to highly precise instruments. But glass
electrode is less subjected to interferences and is used in combination with a calomel
reference electrode. This system is based on the fact that a change of 1 pH unit produces
an electric
charge of 59.1 mV at 25°C.

Turbidity is based on the comparison of the intensity of light scattered by the sample under
defined conditions with the intensity of the light scattered by a standard reference
suspension under the same conditions. The turbidity of the sample is thus measured from
the amount of light scattered by the sample taking a reference with standard turbidity
suspension. The higher the intensity of scattered light the higher is the turbidity. Formazin
polymer is used as the primary standard reference suspension.

Conductivity is a measure of the ability of water to pass an electrical current. Conductivity

in water is affected by the presence of inorganic dissolved solids such as chloride, nitrate,
sulfate, and phosphate anions (ions that carry a negative charge) or sodium, magnesium,
calcium, iron, and aluminum cations (ions that carry a positive charge). Organic
compounds like oil, phenol, alcohol, and sugar do not conduct electrical current very well
and therefore have a low conductivity when in water. Conductivity is also affected by
temperature: the warmer the water, the higher the conductivity. For this reason,
conductivity is reported as conductivity at 25 degrees Celsius (25 0C).

Apparatus:
 Thermometer
 pH meter
 pH paper/pH indicator
 Conductivity meter
 Colorimeter
 Beakers (50 ml)

Procedures:
Sample handling and preservation
Preservation of sample is not practical. Because biological activity will continue after a
sample has been taken, changes may occur during handling and storage. The
characteristics of the water sample may change. To reduce the change in samples taken
for the determination of pH, keep samples at 4°C. Do not allow the samples to freeze.
Analysis should begin as soon as possible.

Precautions
The following precautions should be observed while performing the experiment:
1. Temperature affects the measurement of pH at two points. The first is caused by the
change in electrode output at different temperatures. This interference can be
controlled by the instruments having temperature compensation or by calibrating
the electrode-instrument system at the temperature of the samples. The second is
the change of pH inherent in the sample at different temperatures. This type of error
is sample dependent and cannot be controlled; hence both the pH and
temperature at the time of analysis should be noted.
2. In general, the glass electrode is not subject to solution interferences like color, high
salinity, colloidal matter, oxidants, turbidity or reductants.
3. Oil and grease, if present in the electrode layer, should be removed by gentle
wiping or detergent washing, followed by rinsing with distilled water, because it
could impair the electrode response.
4. Before using, allow the electrode to stand in dilute hydrochloric acid solution for at
least 2 hours.
5. Electrodes used in the pH meter are highly fragile, hence handle it carefully.

1. Determination of the Temperature

1) Wash 50 ml beaker with distilled water.
2) Measure 10 milliliters of water sample, then pour it in the 50 ml beaker.
3) Dip the thermometer in the water sample, and wait until the reading stabilizes.
4) Record your readings.

2. Determination of pH
a) Using pH papers
1) Dip a pH paper from the same water sample used on the determination of
temperature.
2) Compare the color with that of the color given on the wrapper of the pH paper
book.
3) Note down the pH of the sample along with its temperature.
b) Using pH meter
1) Following the manufacturer’s operating instructions of the pH meter, dip the
electrode in the buffer solution of known pH for calibration.
2) Switch on the power supply start the calibration.

3. Determination of turbidity
Note: Follow the manufacturer’s operating instructions of the colorimeter.
1) Calibrate the colorimeter by using a distilled water as a standard for reading
values.
2) Wash the cuvette with distilled water.
3) Fill the cuvette with distilled water, which will be used as a standard-zero reading.
4) Place it on the cuvette holder of the colorimeter and press zero reading.
5) Upon calibration, rinse the cuvette with a small amount of the water sample,
and fill again the cuvette with the water sample.
6) Place it on the cuvette holder of the colorimeter.
7) Press the OK/READ button, and take the reading for turbidity.

4. Determination of conductivity
1) Following the manufacturer’s operating instructions of the conductivity meter,
dip the electrode in distilled water for standardization.
2) Rinse the electrodes with sample water, and then dip it on the same water
sample used on the determination of temperature.
3) Take the conductance reading.

Group Number: _____

Name of Members: ____________________________________
____________________________________
____________________________________
____________________________________
Schedule: ____________________________________

SAMPLE TRIAL TEMPERATURE pH (pH paper) pH (pH meter) TURBIDITY CONDUCTIVITY

1 2

2 2

Note: The following questions should be included in your final report under Answers to
Questions part.

1. Explain why pH is one of the most important controlling factors for treatment and
chemical analysis of water.
2. What is the relationship of pH and Temperature in water?
3. Why is it important to know turbidity in relation to filtration and the disinfection
process in water quality?
4. What are the effects of high and low conductivity in water?

Experiment 2
Determination of Solids

Principle:
Solids analysis provides one of the fundamental measurements used for control of the
activated sludge process and for the regulation of wastewater discharges.

Gravimetric analysis is based on the determination of constituents or categories of

materials by measurement of their weight. The experiment illustrates the principles of
weighing and demonstrates separation and categorization techniques used to define the
various types of solids in waters and wastewaters. These techniques involve three analytic
operations in addition to weighing. These are: filtration, evaporation, and combustion.

Filtration is used to separate suspended or particulate (nonfilterable) fraction from dissolved

or soluble (filterable) fractions. Evaporation separates water from material dissolved or
suspended in it. Combustion differentiates between organic and inorganic matter.
Organic matter will be destroyed completely by burning at 550 0C for 30 min.

Total solids is the term applied to the material left in the vessel after evaporation of a
sample of water/waste water and its subsequent drying in an oven at a definite
temperature. Total solids include “total suspended solids” the portion of total solids retained
by a filter and “total dissolved solids” the portion that passes through the filter. Fixed solids is
the residue remaining after ignition for 1 hour at 550°C. The solid portion that is volatilized
during ignition is called volatile solids. It will be mostly organic matter. Waters that are low in
organic matter and total mineral content and are intended for human consumption may
be examined under 103–105°C or 179–181°C. But water containing considerable organic
matter or those with pH over 9.0 should be dried at 179–181°C. In any case, the report
should indicate the drying temperature. The sample is filtered and the filtrate evaporate in
a weighed dish on a steam bath, the residue left after evaporation is dried to constant
weight in an oven at either 103–105°C or 179–181°C. The increase in weight over that of the
empty dish represents total dissolved solids and includes all materials, liquid or solid, in
solution or otherwise, which passes through the filter and not volatilized during the drying
process.

The difference between the total solids and the total dissolved solids will give the total
suspended solids. The dishes with the residue retained after completion of the tests for total
solids and total dissolved solids are subjected to heat for 1 hour in a muffle furnace held at
550°C. The increase in weight over that of the ignited empty vessel represents fixed solids in
each instance. The difference between the total dissolved/total suspended solids and the
corresponding fixed solids will give volatile solids in each instance. All the quantities should
be expressed in mg/L. Settleable matter in surface and saline waters as well as domestic
and industrial wastes may be determined and reported on a volume basis as milliliter per
liter.

Apparatus:

Procedures:
Sample handling and preservation
Preservation of sample is not practical. Because biological activity will continue after a
sample has been taken, changes may occur during handling and storage. The
characteristics of the water sample may change. To reduce the change in samples taken
for the determination of solids, keep samples at 4°C. Do not allow the samples to freeze.
Analysis should begin as soon as possible.

Precautions
The following precautions should be observed while performing the experiment:
1. Water or Wastewater samples which contain high concentrations of calcium,
chloride, magnesium or sulphate can rapidly absorb moisture from the air. Such
samples may need to be dried for a longer period of time, cooled under proper
desiccation and weighed rapidly in order to achieve a reasonable constant weight.
We should be aware prolonged drying may result in loss of constituents, particularly
nitrates and chlorides.In general, the glass electrode is not subject to solution
interferences like color, high salinity, colloidal matter, oxidants, turbidity or
reductants.
2. Non-representative particulates such as leaves, sticks, fish and lumps of fecal matter
should be excluded from the sample if it is determined that their inclusion is not
desired in the final result.
3. Floating oil and grease, if present, should be included in the sample and dispersed
by a blender device before sub-sampling.
4. Volume of sample should be adjusted to have residue left after drying as 100 to
200mg. It is mainly to prevent large amount of residue in entrapping water during
evaporation.
5. Highly mineralized water containing significant concentration of calcium,
magnesium, chloride, and/or sulphate may be hygroscopic. Hence prolonged
drying, desiccation and rapid weighing.
6. We should be aware prolonged drying may result in loss of constituents, particularly
nitrates and chlorides.

Main Procedure:
1. Label and weigh the filter paper that will be used in the filtration of the sample.
2. Also, label and weigh the crucibles that is to be used in the entire analysis of the
water sample.
3. Set-up the filtration unit.

For Total Solids:

1. Place the crucible (labeled and weight from step 2 in the main procedure) with 25
ml sample in a hot plate and evaporate to dryness.
2. After evaporation of the H2O in the sample, place it in the oven and dry the sample
at a temperature of 1030C to 1050C for 1 hour.
3. After drying, cool in a desiccator to balance temperature and weigh [TS].
4. Repeat step 2 and 3, until a constant weight [TS] is obtained.
5. Place the crucible in a muffle furnace and ignite @ a temperature of 5500C for 15
mins.
6. After igniting, cool in a desiccator to balance temperature and weigh [TFS].
7. Repeat step 5 and 6, until a constant weight [TFS] is obtained.
8. Calculate for the TVS by subtracting the TFS from TS.

For Total Suspended Solids:

1. From step 7 in the main procedure, place the crucible in an oven, and dry at a
temperature of 1030C to 1050C for 1 hour.
2. After drying, cool in a desiccator to balance temperature and weigh the filter paper
[TSS].
3. Repeat step 1 and 2, until a constant weight [TSS] is obtained.
4. Place the crucible with filter paper in a muffle furnace and ignite @ a temperature
of 5500C for 15 mins.
5. After igniting, cool in a desiccator to balance temperature and weigh the filter
paper [FSS].
6. Repeat step 4 and 5, until a constant weight [FSS] is obtained.
7. Calculate for the VSS by subtracting the FSS from TSS.

For Total Dissolved Solids:

1. From step 8 in the main procedure, dry the samples in the crucible at 1800C for 1
hour.
2. After drying, cool in a desiccator to balance temperature and weigh [TDS].
3. Repeat step 1 and 2, until a constant weight [TDS] is obtained.
4. Place the crucible in a muffle furnace and ignite @ a temperature of 5500C for 15
mins.
5. After igniting, cool in a desiccator to balance temperature and weigh [FDS].
6. Repeat step 4 and 5, until a constant weight [FDS] is obtained.
7. Calculate for the VDS by subtracting the FDS from TDS.

(Note: Constant weight is acceptable if the weight change < 4% of the previous weight OR
0.5 mg, whichever is less.)
Calculations:

Total Dissolved Solids

TDS (𝐴 − 𝐵)𝑥 1000
mg total dissolved solids
=
L 𝑠𝑎𝑚𝑝𝑙𝑒 𝑣𝑜𝑙𝑢𝑚𝑒, 𝑚𝐿
where: A = weight of dried residue + dish, mg
B = weight of dish, mg

Total Suspended Solids

(𝐴 − 𝐵)𝑥 1000
mg total suspended solids/L =
𝑠𝑎𝑚𝑝𝑙𝑒 𝑣𝑜𝑙𝑢𝑚𝑒, 𝑚𝐿
where: A = weight of filter + dried residue, mg
B = weight of filter, mg

Group Number: _____

Total Total Volatile Volatile Fixed Fixed

Total
SAMPLE TRIAL Dissolved Suspended Dissolved Suspended Dissolved Suspended
Solids
Solids Solids Solids Solids Solids Solids

Note: The following questions should be included in your final report under Answers to
Questions part.

1. What significant information is furnished by the determination of volatile solids?

2. How can you relate the total solids with the sludge volume index?
3. What are the possible common sources of solids in ground water and surface
water? Explain briefly.
4. Why water is evaporated at 103oC rather than 100oC in assessment of solid of water?

Module/Unit Assessment: Preliminary Exams

To be posted on Google Classroom or any LMS Software to be preferred for the whole
class.

Introduction

Standard Methods for the Examination of Water and Wastewater is the result of a joint
effort by three technical societies: American Public Health Association (APHA), American
Water Works Association (AWWA), and Water Environment Federation (WEF).

The Clean Water Act serves as a requirement nowadays to continually improve the quality
of water and process wastewater effluent discharges. At the same time, population,
production from industries, institutions and commercial entities have increased water use,
creating a corresponding rise in wastewater quantity.

This increased water use and process generations requires more efficient detection and
removal of contaminants in water within the established environmental regulatory limits or
standards.

Experiment 3
Determination of Chlorides

Principle:
Chloride in the form of chloride (Cl-) ion is one of the major inorganic anions in water and
wastewater. The chloride concentration is higher in wastewater than in raw water because
sodium chloride is a common article of diet and passes unchanged through the digestive
system (Average estimate of excretion: 6 g of chlorides/person/day; additional chloride
burden due to human consumption on wastewater: 15 mg/L). Along the sea coast
chloride may be present in high concentration because of leakage of salt water into the
sewage system. It also may be increased by industrial process. In potable water, the salty
taste produced by chloride concentration is variable and depends on the chemical
composition of water. Some waters containing 250 mg/L Cl- may have a detectable salty
taste if sodium cation is present. On the other hand, the typical salty taste may be
absent in waters containing as much as 1000 mg/L when the predominant cations are
calcium and magnesium. In addition, a high chloride contents may harm metallic pipes
and structures as well as growing plants.

The measured chloride ions can be used to know salinity of different water sources. For
brackish water (or sea water or industrial brine solution), it is an important parameter and
indicates the extent of desalting of apparatus required. It also interferes with COD
determination and thus it requires a correction to be made on the basis of amount present
or else a complexing agent, such as HgSO4 can be added. Further, chloride ions are used
as tracer ions in column studies to model fate of different contaminants in soil and liquid
media.

This method determines the chloride ion concentration of a solution by titration with silver
nitrate. As the silver nitrate solution is slowly added, a precipitate of silver chloride forms.
Ag+(aq)+ Cl-(aq) → AgCl(s)

The end point of the titration occurs when all the chloride ions are precipitated. Then
additional silver ions react with the chromate ions of the indicator, potassium chromate, to
form a red-brown precipitate of silver chromate.
2Ag+(aq) + CrO42-(aq)→ Ag2CrO4(s)

This method can be used to determine the chloride ion concentration of water samples
from many sources such as seawater, stream water, river water and estuary water. The pH
of the sample solutions should be between 6.5 and 10. If the solutions are acidic, the
gravimetric method or Volhard’s method should be used.

Apparatus:
 Erlenmeyer flask, 250-ml
 Buret, 50-ml
 Graduated Cylinder

Procedures:
Precautions
The following precautions should be observed while performing the experiment:

1. A uniform sample size must be used, preferably 100 ml (or 50 mL), so that ionic
concentrations needed to indicate the end point will be constant.
2. The pH must be in the range of 7 to 8 because Ag+ is precipitated as AgOH at high
pH levels and the CrO42- is converted to Cr2O72-- at low pH levels,
3. A definite amount of indicator must be used to provide a certain concentration of
CrO4; otherwise Ag2CrO4 may form too soon or not soon enough.
4. The chromate solution needs to be prepared and used with care as chromate is a
known carcinogen.
5. Silver nitrate solution causes staining of skin and fabric (chemical burns). Any spills
should be rinsed with water immediately.

1. Wash the 250-ml Erlenmeyer flask with distilled water then rinse with your water sample.
2. Measure 100 ml of water sample and place it on the Erlenmeyer flask.
3. Adjust sample pH to 7 to 10 with H2SO4 or NaOH if it is not in this range.
4. Add 1.0 mL K2CrO4 indicator solution.
5. Titrate with standard AgNO3 titrant to a pinkish yellow end point. Be consistent in end-
point recognition.

Note: If more than 7 or 8 mL of silver nitrate solution are required, the entire procedure
should be repeated using a smaller sample diluted to 50 ml with distilled water. In here
a blank(distilled water) is needed to determine its Cl- concentration.

Calculations:
For pure sample:
𝑚𝑔 𝐶𝑙 − 𝐴 𝑥 𝑁 𝑥 35,450
=
𝐿 𝑠𝑎𝑚𝑝𝑙𝑒 𝑣𝑜𝑙𝑢𝑚𝑒, 𝑚𝐿
where: A = mL titration for sample

For diluted sample:

𝑚𝑔 𝐶𝑙 − (𝐴 − 𝐵) 𝑥 𝑁 𝑥 35,450
=
𝐿 𝑠𝑎𝑚𝑝𝑙𝑒 𝑣𝑜𝑙𝑢𝑚𝑒, 𝑚𝐿
where: A = mL titration for sample
B = mL titration for blank
N = Normality of AgNO3

𝑚𝑔 𝑁𝑎𝐶𝑙 𝑚𝑔 𝐶𝑙 −
= 𝑥1.65
𝐿 𝐿

Group Number: _____

Standard Silver Nitrate Concentration: _________________

Volume Volume A, ml
B, ml
of of blank titration
SAMPLE TRIAL titration for Cl- mg/L NaCl mg/L
sample used for
blank
used sample

1 2

Note: The following questions should be included in your final report under Answers to
Questions part.

1. How does chlorides being introduced in natural water?

2. Why do we need to have the indicator “blank” or “error” be subtracted from the
amount of silver nitrate used in titiration?
3. What are other methods of determining chlorides in water sample?
4. Why is it important to adjust the pH for the analysis of chlorides?

Experiment 4
Determination of Iron

Principle:
Iron is usually present in natural water and is not objectionable, if concentration is less than
0.3 ppm. It may be in true solution in colloidal state that may be peptized by organic
matter, in the inorganic and organic iron complexes, or in relatively coarse suspended
particles. It may be either ferrous or ferric, suspended or filterable. Iron exists in soils and
minerals mainly as insoluble ferric oxide and iron sulphide (pyrite). It occurs in some areas,
also as ferrous carbonate (siderite), which is very slightly soluble.

Iron may be present in two forms, namely the reduced form (ferrous, Fe2+) and the fully
oxidized form (ferric, Fe3+). Ferric iron is seldom found in true solution in natural waters,
unless they are highly acidic, because of the formation of insoluble ferric hydroxides.
Ferrous iron is more likely to be found in true solution, although it is easily oxidized to the
ferric state and precipitated in alkaline waters as ferric hydroxide.

Since some iron may exist as iron hydroxide precipitates, therefore it is necessary to
bring precipitated form(s) of iron back in 'to solution before oxidizing total iron content
in water. For this hydrochloric acid is added to the test sample to dissolve the insoluble
ferric forms.

For determination of total iron by the following procedure, it must be ensured that all
iron exists in ferric form (Fe3+). This is most readily accomplished by using potassium
permanganate, an oxidizing agent.
5Fe2+ + MnO4- + 8H+ = 5Fe3+ + Mn2+ + 4H2O

Ferric iron is determined by producing a red-colored iron compound, ferric

thiocyanate, by the addition of potassium thiocyanate (Eq.-9.3).
Fe3+ + 3 KCNS = Fe(CNS)3 + 3K+
(red)

The quantity of ferric iron is determined by comparison with the red color produced by
standard iron solutions.

Apparatus:
 Erlenmeyer flask, 250-ml
 Buret, 50-ml
 Micropipette
 Graduated Cylinder
 Rubber Bulb/Pipetol

Reagents:
 Standardized Sulfuric Acid, H2SO4 (3 N)

Procedures:
1. Wash the 250-ml Erlenmeyer flask with distilled water then rinse with your water sample.
2. Measure 25 ml of water sample and place it on the Erlenmeyer flask.
3. Add 8 mL of 3N H2SO4 indicator solution.
4. Titrate with 0.1 N KMnO4 solution to light pink endpoint.

Calculations:

𝑚𝑔 𝑓KMnO4 𝑥 𝑉KMnO4 x 𝑀𝑊𝐹𝑒 x 1000

𝐹𝑒, =
𝐿 𝑓𝐹𝑒 𝑥 𝑚𝐿 𝑠𝑎𝑚𝑝𝑙𝑒

Group Number: _____

Volume of titrant AVERAGE mg

Sample Trials mg Fe/L
used, mL Fe/L

1 2

2 2

Note: The following questions should be included in your final report under Answers to
Questions part.

1. Explain the negative results of Iron in water quality?

2. Is iron considered as a harmful contaminant in water? Explain.
3. What is the effect of high iron content of water in human skin?

Principle:
Hard waters are generally considered to be those waters that require considerable
amounts of soap to produce foam or leather and that also produce scale in hot-water
pipes, boilers, and other units in which the temperature of water is increased substantially.
The hardness of water varies considerably from place to place. In general, surface water is
softer than groundwater. The hardness of water reflects the nature of the geological
formations with which it has been in contact.

Hardness is caused by multivalent metallic cations. Such cations are capable of reacting
with soap to form precipitates and with certain anions present in water to form scale. The
principal hardness causing cations are the divalent calcium, magnesium, strontium, ferrous
ion and manganaous ions. These cations and the important anions with they are
associated with are shown in the following table in the order of their relative abundance in
natural waters. Aluminum and ferric ions are sometimes considered as contributing to the
hardness of water. However, their solubility is so limited at pH values of natural waters that
ionic concentrations are negligible. The hardness of water is derived largely from contact
with the soil and rock formations.

Principal hardness causing cations and the major anions associated with them
Cations causing Associated
Hardness Anions
Ca2+ HCO3-
Mg2+ SO42-
Sr2+ Cl-
Fe2+ NO3-
Mn2+ SiO32-

Classification of hardness types

Degree of
Hardness (mg/L)
Hardness
0 - 75 Soft
Moderately
75 – 100
hard
150 – 300 Hard
> 300 Very Hard

Total hardness is defined as the sum of the calcium and magnesium concentrations, both
expressed as calcium carbonate in mg/L. When hardness numerically is greater than the
sum of carbonate and bicarbonate alkalinity, that amount of hardness equivalent to the
total alkalinity is called “carbonate hardness”; the amount of hardness in excess of this is
called “noncarbonated hardness.” When the hardness numerically is equal to or less than
the sum of carbonate and bicarbonate alkalinity, all hardness is carbonate hardness and
noncarbonate hardness is absent. The hardness may range from zero to hundreds of

Ethylenediaminetetraacetic acid and its sodium salts (abbreviated EDTA) form a chelated
soluble complex when added to a solution of certain metal cations. If a small amount of a
dye such as Eriochrome Black T or Calmagite is added to an aqueous solution containing
calcium and magnesium ions at a pH of 10.0 ± 0.1, the solution becomes wine red. If EDTA
is added as a titrant, the calcium and magnesium will be complexed, and when all of the
magnesium and calcium has been complexed the solution turns from wine red to blue,
marking the end point of the titration. Magnesium ion must be present to yield a
satisfactory end point. To insure this, a small amount of complexometrically neutral
magnesium salt of EDTA is added to the buffer; this automatically introduces sufficient
magnesium and obviates the need for a blank correction. The sharpness of the end point
increases with increasing pH. However, the pH cannot be increased indefinitely because of
the danger of precipitating calcium carbonate, CaCO3, or magnesium hydroxide,
Mg(OH)2, and because the dye changes color at high pH values. The specified pH of 10.0 ±
0.1 is a satisfactory compromise. A limit of 5 min is set for the duration of the titration to
minimize the tendency toward CaCO3 precipitation.

Apparatus:
 Erlenmeyer flask, 250-ml
 Buret, 50-ml
 Graduated Cylinder
 Rubber Bulb/Pipetol

Reagents:
 Buffer Solution (Choose from the two solutions below to which reagents are
available in the chemical counter)
a) Dissolve 16.9 g ammonium chloride (NH4Cl) in 143 mL conc ammonium
hydroxide (NH4OH). Add 1.25 g magnesium salt of EDTA (available commercially)
and dilute to 250 mL with distilled water.
b) Dissolve 1.179 g disodium salt of ethylenediaminetetraacetic acid dihydrate
(analytical reagent grade) and 780 mg magnesium sulfate (MgSO4 ● 7H2O) or
644 mg magnesium chloride (MgCl2 ● 6H2O) in 50 mL distilled water. Add this
solution to 16.9 g NH4Cl and 143 mL conc NH4OH with mixing and dilute to 250 mL
with distilled water. Store in a borosilicate glass container for no longer 1 month.
 Eriochrome Black T
Sodium salt of 1-(1-hydroxy-2-naphthylazo)-5-nitro-2-naphthol-4-sulfonic acid; No.
203 in the Color Index. Dissolve 0.5 g dye in 100 g 2,2’,2”-nitrilotriethanol (also called
triethanolamine) or 2-methoxymethanol (also called ethylene glycol monomethyl
ether). Add 2 drops per 50 mL solution to be titrated. Adjust volume if necessary.
 Standard EDTA titrant, (0.01 M/0.02 N)
Weigh 3.723 g analytical reagent-grade disodium ethylenediaminetetraacetate
dihydrate, also called (ethylenedinitrilo)tetraacetic acid disodium salt (EDTA),
dissolve in distilled water, and dilute to 1000 mL. Standardize against standard
calcium solution
 Standard calcium solution

Procedures:
For Total Hardness
1. Wash the 250-ml Erlenmeyer flask with distilled water then rinse with your water sample.
2. Measure 25 ml of water sample and 25 ml of distilled water and then mix it on the
Erlenmeyer flask.
3. Add 2 mL of buffer solution and 2 drops of indicator solution (Eriochrome Black T).
4. Titrate the sample with standard EDTA titrant until ‘reddish tinge’ color of the sample
disappears.
5. Add the last few drops at 3- to 5-s intervals. At the end point the solution normally is
blue.
6. Record the volume of titrant used for titration to calculate for the total hardness.

For Permanent Hardness

7. Boil 250 ml sample for 15 minutes to oxidize the carbonate ions into carbon dioxide
which will escape from the sample.
8. Repeat steps 1 to 5.
9. Record the volume of titrant used for titration to calculate for the permanent hardness.

Calculations:

𝑚𝑔 𝐶𝑎𝐶𝑂3 𝐴 𝑥 𝐵 𝑥 50 𝑥 1000
𝐻𝑎𝑟𝑑𝑛𝑒𝑠𝑠, =
𝐿 𝑚𝑙 𝑜𝑓 𝑠𝑎𝑚𝑝𝑙𝑒
Where: A = volume of titrant used, ml
B = Normality of EDTA
50 = equivalent weight of CaCO3

Group Number: _____

Standard EDTA titrant Concentration: _________________

Volume of AVERAGE
Volume of Hardness as mg
SAMPLE TRIAL sample used, Hardness as mg
titrant used, ml CaCO3/L
ml CaCO3/L

1 (for TOTAL
2
HARDNESS)

2 (for
PERMANENT 2
HARDNESS)

Note: The following questions should be included in your final report under Answers to
Questions part.

1. What are the effects of hard water in household activities?

2. What are the common methods on how to treat hard water?
3. A sample has 50 mg/L Ca2+, 150 mg/L Mg2+, 50 mg/L Cl- and 100 mg/L glucose.
Calculate the total hardness, carbonate and non-carbonate hardness of water
sample.

Experiment 6
Determination of Nitrates

Principle:
Determination of nitrate (NO3-) is difficult because of the relatively complex procedures
required, the high probability that interfering constituents will be present, and the limited
concentration ranges of the various techniques.

An ultraviolet (UV) technique that measures the absorbance of NO3- at 220 nm is suitable
for screening uncontaminated water (low in organic matter). Screen a sample; if
necessary, then select a method suitable for its concentration range and probable
interferences. Nitrate may be determined by ion chromatography (Section 4110 of APHA),
capillary ion electrophoresis (Section 4140 of APHA), or the methods shown here.
Applicable ranges are: nitrate electrode method, 0.14 to 1400 mg NO3- -N/L; cadmium
reduction method, 0.01 to 1.0 mg NO3- -N/L; automated hydrazine reduction method, 0.01
to 10 mg NO3- -N/L; automated cadmium reduction method, 0.1 to 10 mg NO3- -N/L;
cadmium reduction flow injection method, 0.00025 to 10 mg NO3- -N/L; second-derivative
ultraviolet spectrophotometric method, 0.05 to 2 mg NO3- -N/L. For higher NO3- -N
concentrations, dilute into the range of the selected method.

The colorimetric and UV second-derivative methods require an optically clear sample. Filter
turbid sample through 0.45 μm membrane filter. Test filters for nitrate contamination.

The UV technique is used only for screening samples that have low organic matter
contents, i.e., uncontaminated natural waters and potable water supplies. The NO3-
calibration curve follows Beer’s law up to 11 mg N/L.

Measurement of UV absorption at 220 nm enables rapid determination of NO3-. Because

dissolved organic matter also may absorb at 220 nm and NO3- does not absorb at 275 nm,
a second measurement made at 275 nm may be used to correct the NO3- value. The
extent of this empirical correction is related to the nature and concentration of organic
matter and may vary from one water to another. Consequently, this method is not
recommended if a significant correction for organic matter absorbance is required,
although it may be useful in monitoring NO3- levels within a water body with a constant
type of organic matter. Correction factors for organic matter absorbance can be
established by the method of additions in combination with analysis of the original NO3-
content by another method. Sample filtration is intended to remove possible interference
from suspended particles. Acidification with 1N HCl is designed to prevent interference
from hydroxide or carbonate concentrations up to 1000 mg CaCO3/L. Chloride has no
effect on the determination.

Apparatus:
 Erlenmeyer flask, 250-ml
 Graduated Cylinder
 Rubber Bulb/Pipetol

Reagents:
 Nitrate-free water
Use redistilled or distilled, deionized water of highest purity to prepare all solutions
and dilutions.
 Stock Nitrate solution
Dry potassium nitrate (KNO3) in an oven at 105°C for 24 h. Dissolve 0.7218 g in water
and dilute to 1000 mL; 1.00 mL = 100 μg NO3- -N. Preserve with 2 mL CHCl3/L. This
solution is stable for at least 6 months.
 Intermediate nitrate solution
Dilute 100 mL stock nitrate solution to 1000 mL with water; 1.00 mL = 10 μg NO3- -N.
Preserve with 2 mL CHCl3/L. This solution is stable for at least 6 months.
 Hydrochloric acid solution, HCl (1N)

Procedures:
Preparation of standard curve:
1. Prepare NO3- calibration standards in the range of 0 to 7 mg NO3- -N/L by diluting to 50
mL the following volumes of intermediate nitrate solution: 0, 1.00, 2.00, 4.00, 7.00 . . . 35.0
mL.
2. Treat NO3- standards with 1 ml of HCl solution and mix thoroughly.
3. Proceed to spectrophotometric measurement.

For the water sample:

1. Filter 50 ml of water sample and collect the filtrate in an Erlenmeyer flask.
2. Add 1 mL of HCl solution and mix thoroughly.
3. Proceed to spectrophotometric measurement.

Spectrophotometric measurement:
1. Set the wavelength at 220 nm.
2. Read absorbance or transmittance against redistilled water mixed and set at zero
absorbance or 100% transmittance.
3. Put a sufficient amount of water sample/NO3- standards treated with HCl in the sample
holder (cuvette).
4. Place the cuvette in the cuvette holder of the spectrophotometer.
5. Read for the absorbance or transmittance and record it.
6. Set the wavelength at 275 nm to determine interference due to dissolved organic
matter.
7. Repeat step 2 to 5.

Calculations:
For samples and standards, subtract two times the absorbance reading at 275 nm from the
reading at 220 nm to obtain absorbance due to NO3-. Construct a standard curve by
plotting absorbance due to NO3- against NO3- -N concentration of standard. Using

NOTE: If correction value is more than 10% of the reading at 220 nm, do not use this
method.

Group Number: _____

For calibration curve:

Assigned Group: _____________
Volume of Calculated
Absorbance Absorbance Corrected
intermediate Concentration after
reading @ 220 nm reading @ 275 nm Absorbance
solution, mL dilution mg NO3-/L
1
2
4
7
11
16
22
29
35

For water sample:

Volume
Absorbance Absorbance
of Corrected AVERAGE
SAMPLE TRIAL reading @ 220 reading @ mg NO3-/L
sample Absorbance mg NO3-/L
nm 275 nm
used, ml

1 2

Note: The following questions should be included in your final report under Answers to
Questions part.

1. What are the different forms of nitrogen that normally occurs in water?
2. Discuss the significance of nitrates analysis in water pollution control?
3. What are the resulting environmental occurrences with the presence of nitrites and
nitrates in water?

Principle:
The two major factors that influence selection of the method to determine ammonia are
concentration and presence of interferences. In general, direct manual determination of
low concentrations of ammonia is confined to drinking waters, clean surface or
groundwater, and good-quality nitrified wastewater effluent. In other instances, and where
interferences are present or greater precision is necessary, a preliminary distillation step is
required (See 4500-NH3 B of APHA).

An intensely blue compound, indophenol, is formed by the reaction of ammonia,

hypochlorite, and phenol catalyzed by sodium nitroprusside.

Apparatus:
 Erlenmeyer flask, 50-ml
 Graduated Cylinder
 Rubber Bulb/Pipetol
 Spectrophotometer (UV-Vis)
 Filter Paper
 Funnel

Reagents:
 Phenol solution
Mix 11.1 mL liquified phenol (≥89%) with 95% v/v ethyl alcohol to a final volume of
100 mL. Prepare weekly. CAUTION: Wear gloves and eye protection when handling
phenol; use good ventilation to minimize all personnel exposure to this toxic volatile
substance.
 Sodium nitroprusside, 0.5% w/v
Dissolve 0.5 g sodium nitroprusside in 100 mL deionized water. Store in amber bottle
for up to 1 month.
 Alkaline citrate
Dissolve 200 g trisodium citrate and 10 g sodium hydroxide in deionized water. Dilute
to 1000 mL.
 Sodium hypochlorite
Commercial solution, about 5%. This solution slowly decomposes once the seal on
the bottle cap is broken. Replace about every 2 months.
 Oxidizing solution
Mix 100 mL alkaline citrate solution with 25 mL sodium hypochlorite. Prepare fresh
daily.
 Stock ammonium solution
Dissolve 3.819 g anhydrous NH4Cl (dried at 100°C) in water, and dilute to 1000 mL;
1.00 mL = 1.00 mg N = 1.22 mg NH3.
 Standard ammonium solution
Prepare a series of standard solutions covering the concentrations of 1000 to 0.1 mg
NH3 -N/L by making decimal dilutions of stock NH4Cl solution with water.

For the water sample:

1. Measure 25 ml of water sample and place it in an Erlenmeyer flask.
2. Add 1 mL of phenol solution, 1 ml sodium nitroprusside solution, and 2.5 ml oxidizing
solution and mix thoroughly.
3. Cover samples with plastic wrap or paraffin wrapper film. Let color develop at room
temperature (22 to 27°C) in subdued light for at least 1 hr.
4. Proceed to spectrophotometric measurement.

Spectrophotometric measurement:
1. Set the wavelength at 640 nm.
2. Read absorbance against redistilled water and set at zero absorbance or 100%
transmittance.
3. Put a sufficient amount of water sample/NH3 standards treated with the reagents in the
sample holder (cuvette).
4. Place the cuvette in the cuvette holder of the spectrophotometer.
5. Read for the absorbance or and record it.

Calculations:
Construct a standard curve by plotting absorbance due to NH3 -N against NH3 -N
concentration of standard. Compute sample concentration by comparing sample
absorbance with the standard curve.

Group Number: _____

For calibration curve:

Assigned Group: _____________
Volume of stock Calculated
Absorbance
ammonium solution, Concentration after
reading @ 640 nm
mL dilution mg NH3 -N/L
0.025
.125
.25
1.25
2.5
12.5
Pure stock solution

For water sample:

Volume
Absorbance AVERAGE
of mg NH3 -
SAMPLE TRIAL reading @ 640 mg NH3 -
sample N/L
nm N/L
used, ml

1 2

Note: The following questions should be included in your final report under Answers to
Questions part.

1. How does ammonia and chlorine be related in terms of water quality?

2. What is the range of acceptable concentration of ammonia in water? Explain
briefly.
3. What other methods can be used to determine ammonia in water?

Experiment 8
Determination of Sulfates

Principle:
Sulfate are found in appreciable quantity in all natural waters, particularly high in arid and
semi arid regions where natural waters in general have high salt content. Sulfate salts are
mostly soluble in water and impart hardness. Water with high concentrations has a bitter
test. Sulfate may cause intestinal disorders. These ions can produce hydrogen sulfides as
per following equation (1):
SO42- + organic matter → S2+ + H2O + CO2 (in the presence of anaerobic bacteria)
S2- + H+ ↔ HS-
HS- + H+ ↔ H2S

The sulfate data is used in determining applicability of different water types for their public
and industrial applications. It indirectly indicates extent of problems that can arise due to
reduction of sulfates to hydrogen sulfides. In addition, sulfate content of organic matter fed
to anaerobic digester is important information as it gives idea of generation of hydrogen
sulfides, which needs to be removed.

Sulfate ion (SO42-) is precipitated in an acetic acid medium with barium chloride (BaCl2) so
as to form barium sulfate (BaSO4) crystals of uniform size. Light absorbance of the BaSO4
suspension is measured by a photometer and the SO42- concentration is determined by
comparison of the reading with a standard curve.

Apparatus:
 Erlenmeyer flask, 250-ml
 Graduated Cylinder
 Rubber Bulb/Pipetol
 Spectrophotometer (UV-Vis)
 Stopwatch

Reagents:
 Buffer solution A
Dissolve 30 g of magnesium chloride, MgCl2.6H2O, 5 g sodium acetate,
CH3COONa.3H2O, 1.0 g potassium nitrate, KNO3, and 20 mL acetic acid, CH3COOH
(99%), in 500 ml distilled water and added water until the solution is 1000mL.
 Buffer solution B (required when the sample SO42- concentration is less than 10 mg/L)
Dissolve 30 g of magnesium chloride, MgCl2.6H2O, 5 g sodium acetate,
CH3COONa.3H2O, 1.0 g potassium nitrate, KNO3, 0.111 g sodium sulfate and 20 mL
acetic acid, CH3COOH (99%), in 500 ml distilled water and added water until the
solution is 1000mL.
 Barium chloride (20 to 30 mesh)
 Standard sulfate solution

Preparation of standard curve:

1. Prepare SO42- calibration standards in the range of 0 to 40 mg SO42-/L range with space
standards 5 mg/L increments and diluting it to 100 ml.
2. Proceed to spectrophotometric measurement.

For the water sample:

1. Measure 100 ml of water sample and place it in an Erlenmeyer flask.
2. Add 20 mL of buffer solution and mix.
3. While stirring, add a spoonful of BaCl2 crystals and begin timing immediately. Stir for 60 ±
2 s at constant speed.
4. Proceed to spectrophotometric measurement.

Correction for sample color and turbidity of the water sample:

5. Measure 100 ml of water sample and place it in an Erlenmeyer flask.
6. Add 20 mL of buffer solution and mix.
7. Proceed to spectrophotometric measurement.

Spectrophotometric measurement:
6. Set the wavelength at 420 nm.
7. Read absorbance against redistilled water and set at zero absorbance or 100%
transmittance.
8. Put a sufficient amount of water sample/SO42- standards treated with the reagents in
the sample holder (cuvette).
9. Place the cuvette in the cuvette holder of the spectrophotometer.
10. Read for the absorbance or and record it.

Calculations:
If buffer solution A was used:
Determine SO42- concentration directly from the calibration curve after subtracting sample
absorbance before adding BaCl2.

If buffer solution B was used:

𝑚𝑔 𝑆𝑂42− 𝑚𝑔 𝑆𝑂42− 𝑥 1000
=
𝐿 𝑚𝐿 𝑠𝑎𝑚𝑝𝑙𝑒
Subtract SO42- concentration of blank from apparent SO42- concentration as determined
above; because the calibration curve is not a straight line, this is not equivalent to
subtracting blank absorbance from sample absorbance.

Group Number: _____

For calibration curve:

Assigned Group: _____________
Concentration, mg Volume of 0.02 N Absorbance
SO42-/L H2SO4 used reading @ 420 nm
0
5
10
15
20
25
30
35
40

For water sample:

Absorbance Absorbance
Volume
reading for reading for
of AVERAGE
sample sample Calculated Concentration,
SAMPLE TRIAL sample Concentration,
treated with untreated Absorbance mg SO42-/L
used, mg SO42-/L
BaCl2 @ 420 with BaCl2 @
ml
nm 420 nm

1 2

Note: The following questions should be included in your final report under Answers to
Questions part.

1. What is the effect of the reduction of sulfates in water?

2. Why is it important to determine sulfates in the design of anaerobic digesters?

Experiment 9
Determination of Phosphates

Principle:
Phosphorus occurs in natural waters and in wastewaters almost solely as phosphates. These
are classified as orthophosphates, condensed phosphates (pyro-, meta-, and other
polyphosphates), and organically bound phosphates. They occur in solution, in particles or
detritus, or in the bodies of aquatic organisms.

These forms of phosphate arise from a variety of sources. Small amounts of orthophosphate
or certain condensed phosphates are added to some water supplies during treatment.
Larger quantities of the same compounds may be added during laundering or other
cleaning, because these materials are major constituents of many commercial cleaning
preparations. Phosphates are used extensively in the treatment of boiler waters.
Orthophosphates applied to agricultural or residential cultivated land as fertilizers are
carried into surface waters with storm runoff and to a lesser extent with melting snow.
Organic phosphates are formed primarily by biological processes. They are contributed to
sewage by body wastes and food residues, and also may be formed from
orthophosphates in biological treatment processes or by receiving-water biota.

Phosphorus is essential to the growth of organisms and can be the nutrient that limits the
primary productivity of a body of water. In instances where phosphate is a growth-limiting
nutrient, the discharge of raw or treated wastewater, agricultural drainage, or certain
industrial wastes to that water may stimulate the growth of photosynthetic aquatic micro-
and macroorganisms in nuisance quantities.

Phosphates also occur in bottom sediments and in biological sludges, both as precipitated
inorganic forms and incorporated into organic compounds.

Ammonium molybdate and antimony potassium tartrate react in acid medium with
orthophosphate to form a heteropoly acid—phosphomolybdic acid—that is reduced to
intensely colored molybdenum blue by ascorbic acid.

Apparatus:
 Erlenmeyer flask, 125-ml
 Graduated Cylinder
 Rubber Bulb/Pipetol
 Spectrophotometer (UV-Vis)
 Stopwatch

Reagents:
 Sulfuric Acid, H2SO4, (5N)
Dilute 70 mL conc H2SO4 to 500 mL with distilled water.
 Antimony potassium tartrate solution

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means, electronic, mechanical, photocopying, recording, or otherwise of any part of this document, without the prior written permission of SLU, is strictly prohibited. 60
Dissolve 1.3715 g K(SbO)C4H4O6 ● 1⁄2 H2O in 400 mL distilled water in a 500-mL
volumetric flask and dilute to volume. Store in a glass-stoppered bottle.
 Ammonium Molybdate solution
Dissolve 20 g (NH4)6Mo7O24 ● 4 H2O in 500 mL distilled water. Store in a
glass-stoppered bottle.
 Ascorbic Acid (0.1 M)
Dissolve 1.76 g of ascorbic acid in 100 mL distilled water. The solution is stable for
about 1 week at 4oC.
 Combined reagent
Mix the above reagents in the following proportions for 100 mL of the combined
reagent: 50 mL 5N H2SO4 , 5 mL antimony potassium tartrate solution, 15 mL
ammonium molybdate solution, and 30 mL ascorbic acid solution. Mix after addition
of each reagent. Let all reagents reach room temperature before they are mixed
and mix in the order given. If turbidity forms in the combined reagent, shake and let
stand for a few minutes until turbidity disappears before proceeding. The reagent is
stable for 4 h.
 Stock phosphate solution
Dissolve in distilled water 219.5 mg anhydrous KH2PO4 and dilute to 1000 mL; 1.00 mL
= 50.0 μg PO43- -P.
 Standard phosphate solution
Dilute 50.0 mL stock phosphate solution to 1000 mL with distilled water; 1.00 mL = 2.5
μg P.

Procedure:
Preparation of standard curve:
1. Prepare PO43- calibration standards of blank, 0.15, 0.2, 0.5, 1.0, and 1.3 mg PO43- -P/L by
diluting the standard phosphate solution to 50 ml.
2. Add 0.05 mL (1 drop) phenolphthalein indicator. If a red color develops, add 5N H2SO4
solution dropwise to just discharge the color.
3. Add 8.0 mL combined reagent and mix thoroughly.
4. Continue mixing for 12 minutes.
5. Proceed to spectrophotometric measurement.

For the water sample:

1. Measure 50 ml of water sample and place it in an Erlenmeyer flask.
2. Add 0.05 mL (1 drop) phenolphthalein indicator. If a red color develops, add 5N H2SO4
solution dropwise to just discharge the color.
3. Add 8.0 mL combined reagent and mix thoroughly.
4. Continue mixing for 12 minutes.
5. Proceed to spectrophotometric measurement.

Correction for sample color and turbidity of the water sample (high colored):
1. Measure 100 ml of water sample and place it in an Erlenmeyer flask.
2. Add 0.05 mL (1 drop) phenolphthalein indicator. If a red color develops, add 5N H2SO4
solution dropwise to just discharge the color.

Spectrophotometric measurement:
1. Set the wavelength at 880 nm.
2. Read absorbance against redistilled water with the combined reagent and set at zero
absorbance or 100% transmittance.
3. Put a sufficient amount of water sample/PO43- standards treated with the reagents in
the sample holder (cuvette).
4. Place the cuvette in the cuvette holder of the spectrophotometer.
5. Read for the absorbance or and record it.

Calculations:
Construct a standard curve by plotting absorbance due to PO43- against PO43-
concentration of standard. Compute sample concentration by comparing sample
absorbance with the standard curve.

Group Number: _____

For calibration curve:

Assigned Group: _____________
Volume of stock
Concentration, mg Absorbance
phosphate solution
P/L reading @ 880 nm
used, mL
0
0.15
0.20
0.50
1.0
1.3

For water sample:

Volume
Absorbance
of AVERAGE
reading for Concentration,
SAMPLE TRIAL sample Concentration,
sample @ mg P/L
used, mg P/L
880 nm
ml

1 2

Note: The following questions should be included in your final report under Answers to
Questions part.

1. Explain how does phosphates contributes in soil erosion in river banks?

2. How does phosphates affect water quality?
3. What is the normal phosphate level in water?

Module/Unit Assessment: Midterm Exams

To be posted on Google Classroom or any LMS Software to be preferred for the whole
class.

Experiment 10
5-Day BOD Test

Principle:
The biochemical oxygen demand determination is a chemical procedure for determining
the amount of dissolved oxygen needed by aerobic organisms in a water body to break
the organic materials present in the given water sample at certain temperature over a
specific period of time.

BOD of water or polluted water is the amount of oxygen required for the biological
decomposition of dissolved organic matter to occur under standard condition at a
standardized time and temperature. Usually, the time is taken as 5 days and the
temperature is 20°C.

The test measures the molecular oxygen utilized during a specified incubation period for
the biochemical degradation of organic material (carbonaceous demand) and the
oxygen used to oxidize inorganic material such as sulfides and ferrous ion. It also may
measure the amount of oxygen used to oxidize reduced forms of nitrogen (nitrogenous
demand).

BOD is the principle test to give an idea of the biodegradability of any sample and strength
of the waste. Hence the amount of pollution can be easily measured by it. Efficiency of
any treatment plant can be judged by considering influent BOD and the effluent BOD and
so also the organic loading on the unit.

Application of the test to organic waste discharges allows calculation of the effect of the
discharges on the oxygen resources of the receiving water. Data from BOD tests are used
for the development of engineering criteria for the design of wastewater treatment plants.
Ordinary domestic sewage may have a BOD of 200 mg/L. Any effluent to be discharged
into natural bodies of water should have BOD less than 30 mg/L. This is important
parameter to assess the pollution of surface waters and ground waters where
contamination occurred due to disposal of domestic and industrial effluents. Drinking water
usually has a BOD of less than 1 mg/L. But, when BOD value reaches 5 mg/L, the water is
doubtful in purity. The determination of BOD is used in studies to measure the self-
purification capacity of streams and serves regulatory authorities as a means of checking
on the quality of effluents discharged to stream waters.

The determination of the BOD of wastes is useful in the design of treatment facilities. It is the
only parameter, to give an idea of the biodegradability of any sample and self-purification
capacity of rivers and streams. The BOD test is among the most important method in
sanitary analysis to determine the polluting power, or strength of sewage, industrial wastes
or polluted water. It serves as a measure of the amount of clean diluting water required for
the successful disposal of sewage by dilution.

Since the oxygen demand of typical waste is sever hundred milligrams per liter, and since
the saturated value of DO for water at 20uC is only 9.1 mg/L, it is usually necessary to dilute
the sample to keep final DO above zero. If during the five days of experiment, the DO
drops to zero, then the test is invalid since more oxygen would have been removed had
more been available.
The five-day BOD of a diluted sample is given by,
𝑩𝑶𝑫𝟓 = [𝑫𝑶𝒊 − 𝑫𝑶𝒇 ] 𝒙 𝑫. 𝑭.
Here,
(𝑽𝒐𝒍𝒖𝒎𝒆 𝒐𝒇 𝒘𝒂𝒔𝒕𝒆𝒘𝒂𝒕𝒆𝒓 + 𝑽𝒐𝒍𝒖𝒎𝒆 𝒐𝒇 𝒅𝒊𝒍𝒖𝒕𝒊𝒐𝒏 𝒘𝒂𝒕𝒆𝒓)
𝑫. 𝑭. =
𝑽𝒐𝒍𝒖𝒎𝒆 𝒐𝒇 𝒘𝒂𝒔𝒕𝒆𝒘𝒂𝒕𝒆𝒓
In some cases, it becomes necessary to seed the dilution water with microorganisms to
ensure that there is an adequate bacterial population to carry out the biodegradation. In
such cases, two sets of BOD bottles must be prepared, one for just the seeded dilution
water (called the "blank") and the other for the mixture of wastewater and dilution wader.
The changes in DO in both are measured. The oxygen demand of waste water (BODw) is
then determined from the following relationship:
𝑩𝑶𝑫𝒎 𝒙 𝑽𝒎 = (𝑩𝑶𝑫𝒘 𝒙 𝑽𝒘 ) + ((𝑩𝑶𝑫𝒅 𝒙 𝑽𝒅 )
Where, BODm, is the BOD of the mixture of wastewater and dilution water and BODd is the
BOD of the dilution water alone; Vw and Vd are the volumes of wastewater and dilution
water respectively in the mixture and Vm = Vw + Vd.

Apparatus:
 BOD Bottle
 Beaker, 1000-ml
 Graduated Cylinder
 Pipet
 Rubber Bulb/Pipetol
 Buret, 50 mL
 Dropper
 Stirrer
 Incubator

Reagents:
For Dissolved Oxygen
 Manganous sulfate solution
Dissolve 480 g MnSO4.4H2O, 400 g MnSO4.2H2O or 364 g MnSO4.H2O in distilled water,
filter, and dilute to 1L. The MnSO4 solution should not give a color with starch when
added to an acidified potassium iodide (KI) solution.
 Alkali-iodide-azide reagent
1. For saturated or less-than-saturated samples – Dissolve 500 g NaOh (or 700 g
KOH) and 135 g NaI (or 150 g KI) in distilled water and dilute to 1 L. Add 10 g
NaN3 dissolved in 40 mL distilled water. Potassium and sodium salts may be

Property of and for the exclusive use of SLU. Reproduction, storing in a retrieval system, distributing, uploading or posting online, or transmitting in any form or by any
means, electronic, mechanical, photocopying, recording, or otherwise of any part of this document, without the prior written permission of SLU, is strictly prohibited. 66
used interchangeably. This reagent should not give a color with starch
solution when diluted and acidified.
2. For supersaturated samples – Dissolve 10 g NaN3 in 500 mL distilled water. Add
480 g sodium hydroxide (NaOH) and 750 g sodium iodide (NaI), and stir unti
dissolved. There will be a white turbidity due to some sodium carbonate
(Na2CO3), but this will do no harm. CAUTION-Do not acidify this solution
because toxic hydrazoic acid fumes may be produced.
 Sulfuric Acid, H2SO4
One mL is equivalent to about 3mL alkali-iodide-azide reagent.
 Starch Solution
Dissolve 2 g laboratory-grade soluble starch and 0.2 g salicyclic acid as preservative
in 100 mL hot distilled water.
 Standard sodium thiosulfate titrant
Dissolve 6.205 g Na2S2O3 .5H2O in distiller water and add 1.5 mL 6N NaOH or 0.4 g
solid NaOH and dilute to 1000 mL. Standardize with bi-iodate solution.
 Standard potassium bi-iodate solution (0.0021 M)
Dissolve 812.4 mg KH(IO3) in distilled water and dilute to 1000 mL.
 Standardization
Dissolve approximately 2 g KI, free from iodate in an Erlenmeyer flask with 100 to 150
mL distilled water; add 1 mL 6N H2SO4 or a few drops of conc. H2SO4 and 20.00 mL
standard bi-iodate solution. Dilute to 200 mL and titrate librated iodine with
thiosulfate titrant, adding starch toward end of titration, when a pale straw color is
reached. When the solution is of equal, 20.00 mL 0.025M Na2S2O3 should be required.
If not, adjust the Na2S2O3 solution to 0.025M.

For BOD-Dilution water

 Phosphate buffer solution
Dissolve 8.5 g KH2PO4, 21.75 g K2HPO4. 33.4 g Na2HPO4 ● 7H2O, and 1.7 g NH4Cl in
about 500 mL distilled water and dilute to 1 L. The pH should be 7.2 without further
adjustment. Alternatively, dissolve 42.5 g KH2PO4 or 54.3 g K2HPO4 in about 700 m
distilled water. Adjust pH to 7.2 with 30% NaOH and dilute to 1 L.
 Magnesium sulfate solution
Dissolve 22.5 g MgSO4 ● 7H2O in distilled water and dilute to 1 L.
 Calcium chloride solution
Dissolve 27.5 g CaCl2 in distilled water and dilute to 1 L.
 Ferric chloride solution
Dissolve 0.25 g FeCl3 ● 6H2O in distilled water and dilute to 1 L.
 Demineralized, distilled, tap, or neutral water

Procedure:
Preparation of Dilution Water:
Note: Observe the dissolved oxygen of a pure water sample first. The resulting dissolved
oxygen of the pure water sample dictates whether or not the samples need to be diluted
or not with dilution water. If the dissolved oxygen of the water sample is quite low, dilution

Initial DO determination
1. Measure 298 mL of water sample and place it on the BOD bottle.
2. Add 1 mL MnSO4 solution, followed by 1 mL alkali-iodide-azide reagent. If pipets are
dipped into sample, rinse them before returning them to the reagent bottles.
Alternatively, hold pipet tips just above liquid surface when adding reagents.
3. Mix carefully by using the pipet.
4. After 2 minutes, add 1 mL of concentrated H2SO4 and mix several times using pipet until
dissolution is complete.
5. Mix it well again for 2 minutes.
6. Measure 200 mL from the BOD bottle and place it immediately in a cleaned Erlenmeyer
flask.
7. Titrate with 0.025 M Na2S2O3 solution to a pale straw color.
8. Add a few drops of starch solution and continue titration to first disappearance of blue
color.

Final DO determination
1. For the water sample with dilution water, measure 100 mL of sample water and 198 mL
of dilution water and place it on the BOD bottles.
2. Mix carefully by using the pipet.
3. Stopper carefully the bottle.
4. Place the BOD bottles with sample and incubate @ 20oC for 5 days in an incubator.
5. After 5 days, take the BOD bottles out of the incubator, and pipet off 2 mL of sample to
before the addition of 1 mL each of MnSO4 and alkali-iodide azide reagent.
6. Add 1 mL MnSO4 solution, followed by 1 mL alkali-iodide-azide reagent. If pipets are
dipped into sample, rinse them before returning them to the reagent bottles.
Alternatively, hold pipet tips just above liquid surface when adding reagents.
7. After 2 minutes, add 1 mL of concentrated H2SO4 and mix several times using pipet until
dissolution is complete.
8. Mix it well again for 2 minutes.
9. Measure 200 mL from the BOD bottle and place it immediately in a cleaned Erlenmeyer
flask.
10. Titrate with 0.025 M Na2S2O3 solution to a pale straw color.
11. Add a few drops of starch solution and continue titration to first disappearance of blue
color.

Calculations:
1 mL of 0.025 M Na2S2O3 = 1 mg DO/L

𝐷𝑂𝑖 − 𝐷𝑂𝑓
𝐵𝑂𝐷5 =
𝑉𝑜𝑙𝑢𝑚𝑒 𝑜𝑓 𝑠𝑎𝑚𝑝𝑙𝑒
𝑉𝑜𝑙𝑢𝑚𝑒 𝑜𝑓 𝑡ℎ𝑒 𝐵𝑂𝐷 𝑏𝑜𝑡𝑡𝑙𝑒

Group Number: _____

Before incubation After incubation

SAMPLE TRIAL
Volume of Volume Volume of Volume of
sample of titrant DOi, mg/L sample titrant used, DOf, mg/L
used, ml used, mL used, ml mL

Note: The following questions should be included in your final report under Answers to
Questions part.

1. What importance will it give by having a dilution in a water sample?

2. In a BOD test on a diluted wastewater sample (1:20 dilution, but not seeded), the
initial DO is 8.2 mg/L and final DO after 5 days is 3.2 mg/L. If the reaction rate
constant is 0.2/day, calculate a) 5-day BOD, b) Ultimate carbonaceous BOD, c)
remaining oxygen demand after 5 days.
3. Give 3 methods to control nitrification in the 5 days BOD test at 20oC?
4. If 5-day BOD at 20oC is equal to 150 mg/L (rate constant, k=0.23/day), calculate the
5-day BOD at 15oC and 22oC. Use the relationship of k with temperature given in
your Environmental Engineering Lecture Class.

Experiment 12
Total Coliform and Fecal Coliform Test

Principle:
Coliform group comprises of all the aerobic, facultative and anaerobic gram-negative
non-spore forming rod shaped bacteria that ferment lactose with gas formation within 48
hours at 35°C. The standard test for this group may be carried out either by multiple tube
fermentation technique or by membrane filter technique. The E.coli test by multiple tube
fermentation technique consists of 3 phases – presumptive, confirmed and completed.

Escherichia coli (E.coli) for the purpose of sanitary examination of water, is defined as a
gram-negative, nonspore forming rod which is capable of fermenting lactose with the
production of acid and gas at 35°C in less than 48 hours, which produces indole peptone
water containing tryptophan, which is incapable of utilising sodium citrate as its sole source
of carbon, which is incapable of producing acetyl methyl carbinol, and which gives a
positive methyl red test. The results are expressed in terms of MPN (Most Probable Number),
which is based on certain probability formulae. The estimate may give a value greater
than the actual number of coliform present. The accuracy of any single test depends on
the number of tubes fermented. This method helps in describing the sanitary quality of
water.

The safety of the water is generally judged from the knowledge of sanitary condition and
mentioned by the number of samples yielding positive or negative results. If more than 95%
should yield negative results, the safety is usually assured. The scheme of the MPN test is
given as follows:

Apparatus:
 Erlenmeyer flask
 Beaker, 1000-ml
 Durham Tube
 Pipet
 Rubber Bulb/Pipetol
 Bunsen Burner
 Culture tube
 Autoclave
 Incubator

Reagents:
For Dissolved Oxygen
 Lauryl Tryptose broth (LSB)
 Brilliant Green Bile Lactose Broth (BGLB)
 Escherichia Cole Broth (ECB)

Procedure:
For Presumptive Test (5 samples)

For Confirmative Test

Fecal Coliform:
1. Measure 10 ml of ECB and put it on a new set of culture tubes.
2. From the positive results of presumptive test, add 2 drops of the solution and put it on
the culture tube.
3. Gently swirled the culture tube to distribute the sample uniformly throughout the
medium and make sure that there are no bubbles inside it.
4. Place the culture tubes in an incubator at 44oC for 24 hours.
5. After 24 hours, examined the tubes for the presence of gas by determining if there are
bubbles formed.
6. Tubes having positive presence of gas are to proceed in the total coliform and fecal
coliform analysis, take note of the number of tubes. Tubes having negative presence of
gas should be put back to the incubator and incubate again for 24 hours.

Total Coliform:
1. Measure 10 ml of BGLB and put it on a new set of culture tubes.
2. From the positive results of presumptive test, add 2 drops of the solution and put it on
the culture tube.
3. Gently swirled the culture tube to distribute the sample uniformly throughout the
medium and make sure that there are no bubbles inside it.
4. Place the culture tubes in an incubator at 44oC for 24 hours.
5. After 24 hours, examined the tubes for the presence of gas by determining if there are
bubbles formed.
6. Tubes having positive presence of gas are to proceed in the total coliform and fecal
coliform analysis, take note of the number of tubes. Tubes having negative presence of
gas should be put back to the incubator and incubate again for 24 hours.

Calculations:

𝑀𝑃𝑁 𝑛𝑜. 𝑜𝑓 𝑝𝑜𝑠𝑖𝑡𝑖𝑣𝑒 𝑡𝑢𝑏𝑒𝑠 ∗ 100

=
100 √𝑚𝑙 𝑜𝑓 𝑠𝑎𝑚𝑝𝑙𝑒 𝑖𝑛 𝑛𝑒𝑔𝑎𝑡𝑖𝑣𝑒 𝑡𝑢𝑏𝑒𝑠 ∗ 𝑚𝑙 𝑜𝑓 𝑠𝑎𝑚𝑝𝑙𝑒 𝑖𝑛 𝑎𝑙𝑙 𝑡𝑢𝑏𝑒𝑠

Group Number: _____

Presumptive Test Fecal Coliform Total Coliform

Number of Number of Number of
MPN/100 MPN/100 MPN/100
Sample positive positive positive
mL mL mL
tubes tubes tubes

Note: The following questions should be included in your final report under Answers to
Questions part.

1. Explain the terms “indicator organism” and “coliform index”.

2. Why water samples are usually tested for indicator organisms instead of specific
pathogenic organisms?
3. Differentiate total coliform from fecal coliform.
4. Explain the major advantage of “membrane filtration method” over “multiple tube
method”.

Module/Unit Assessment: Final Exams (Comprehensive)

To be posted on Google Classroom or any LMS Software to be preferred for the whole
class.

Capstone:
Make a summary of all paramaters determined in the entire semester, and make a general
conclusion based from the average results of water quality testing.

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