The Biological, Chemical, and Physical Condition of the Calumpang

River: Basis for Environmental Program

A Scientific Research Paper

Presented to the Faculty of

STONYHURST SOUTHVILLE INTERNATIONAL SCHOOL

Batangas Campus

In Partial Fulfillment of the Requirements

for Science 10

Sulit, Paul Anthony

May 2020
 CHAPTER I

THE PROBLEM AND ITS BACKGROUND

INTRODUCTION

Water is one of the most essential things that we use in our lives. 

It’s a factor that is being repeatedly used by people to do simple tasks

such as bathing the body, washing of clothes, hydrating one’s body, and

cooking food. It’s also used in a major task such as producing energy. It is

the core producer of hydroelectric energy. It also majorly composes the

Earth itself. However, its quality has declined due to an event called water

pollution.

According to Black (2018), water pollution has negatively impacted

the environment and living things. Human health is affected by it which

results in the rise of waterborne diseases such as cholera and typhoid

fever. The ecosystem is also in danger because the pollution breeds algae

growth and causes the growth of many water organisms. It continuously

grows and attacks fish and other marine life by reducing their oxygen

supply. It causes the death of marine life because of the deadly

components that it's composed of. Finally, it has an impact on economic

cost. The process and purification of the water would cost an expensive

amount of money. The fishing stock would decline because of the

depletion of oxygen. This would also make customers wary of buying fish

or any marine creatures because of the fear of attaining diseases from it.

The Philippines is considered to be one of the highest contributors to

water pollution due to the number of waste products being contributed by

consumers. The country doesn’t have a sustainable waste system that

could deal with those waste and it affects the land and bodies of water. An

example would be the Calumpang River. According to Cinco (2014), the

river is a 20-kilometer body of water that is connected to two different cities

and six towns. The Calumpang River has amassed a high coliform count.

When compared to the DENR standard, the river has to stay below 400

mpn/ ml. All the other indicators of the river haven’t met the standard of the

DENR.

The researcher aimed to provide a detailed analysis of the variable

and a possible solution. This would provide background knowledge on the

quality of Calumpang River water and a potential way for it to match up to

standards. The reason would be the researcher would want to find a way

to sustain and improve the condition of the river for the benefit of the

residents and the government. The researcher’s observation of the river

would be that it is infected. The basis there would be the sewage water

being placed on the river and the presence of material waste on the
surface. If that factor cannot be changed then it could result in health

hazards not only to humans but also to other marine life.

RESEARCH QUESTIONS

This study aimed to find the biological, chemical, and physical

condition of the Calumpang River. It was analyzed and compared to the

standard quality of water which led to the proposed action plan.

Specifically, it aimed to answer the following questions:

1. What is the biological condition of the Calumpang river in terms of:

a. Biological Oxygen Demand;

b. Fecal Matter?

2. What is the chemical condition of the Calumpang River in terms of:

a. pH;

b. Salinity;

c. Dissolved Oxygen?

3. What is the physical condition of the Calumpang River in terms of:

a. Temperature;

b. Color;

c. Total Suspended solids?

 4. Are the biological, chemical, and physical condition of the

Calumpang River meet the standard quality of water?

5. Based on the results, what environmental program could be

proposed?

SCOPE, LIMITATION AND DELIMITATION

This study focuses on the biological, chemical, and physical

condition of the Calumpang River. This would also compare the quality of

the Calumpang River to the DENR no. 2016-06 standard quality of

freshwater. The researcher, along with the staff of the Provincial

Government Environment and Natural Resources Office, Batangas

Environment Laboratory, did three trials within the two stations along the

Calumpang River.

The study is limited to biological conditions which included biological

oxygen demand, and fecal matter; chemical conditions such as pH level,

salinity, and dissolved oxygen; and the physical conditions delimited to

temperature, color, and total suspended solids. The generalization and

inferences are limited to the water samples taken from the Calumpang

River in the area of Batangas City. The samples were taken from Sabang

Ibaan (Upper part of Calumpang River) and Brgy. Wawa(Lower part of

Calumpang River). The researcher will utilize the standard quality of water

from  DENR no. 2016-06 to check if the results meet up with the standard.

SIGNIFICANCE OF THE STUDY

The research will benefit the following groups:

 Fishermen- The fishermen will benefit from this because the river

could be able to support marine life. This opportunity could make

them capture fish or other marine life in that area.

 Residents along the river- The residents will benefit from this

because they could be able to use the river with whatever activity

they want without the worry if it will cause them harm.

 Batangas Government- This research will prove beneficial to them

as they could implement programs, or actions that could clean not

only the Calumpang River  but also other rivers that contain deadly

components.  

 Future Researchers-  The study could be used as a related study

on the research chosen by the future researcher. They could use

this as evidence that supports his or her study.

Conceptual Literature

The researcher discussed the biological, chemical and physical

condition of the Calumpang River. Each subtopic of the three conditions

was discussed in this section to provide a basis of the experiment.

Biological Condition is a major indicator of a body of water for it

shows if one can be able to support living organisms. According to

Kompasiana (2015), the condition means the presence of microorganisms

and organisms living inside the area. An example would be bacteria,

protozoan, insects. and big animals. In this study, it has two subtopics

which include: Biological Oxygen Demand and Fecal Matter.

According to Li (2019), Biological Oxygen Demand (BOD) is the

amount of dissolved oxygen needed by aerobic biological organisms

present in the water. It is commonly expressed in milligrams of oxygen per

liter. It helps show the presence of organic matter for its attendance shows

the appearance of BOD. If there is a high BOD then the amount of

dissolved oxygen will be lesser for the aquatic life to consume. It is also

one of the main reasons for treating water because it could lessen the

demand for streams and rivers. 

 Based on Sharma (2016), Biological Oxygen Demand is several

oxygen molecules needed for the organisms to decay waste. The source

states numerous reasons for its importance. It claims that this is an

important way to test if a body of water is infected. Another importance

would be that most of the results, when tested, are used to create

wastewater treatment plants. Finally, it is one of the factors used to

determine the diluted water needed for the removal of unneeded

substances. The informant states that if the BOD value reaches 5 mg/L

then it can be assumed that the sample is impure.

According to Newal et. al (2015), Fecal Matter is a solid excretory

product that comes from the bowels. It is a waste product of humans, and

animals. It’s important to analyze it because it indicates the shift in water

flow and contamination. They are present in humid areas however it also

shows up in bodies of water thanks to the sewage systems. They linked

that human activity and changes in the environment increase the amount

of fecal matter which leads to the breach in pure water.

 Likewise, the CDC (2017), the germs that cause recreational water

illnesses could be spread through contaminated water thanks to fecal

matter. The organization goes on to say that they could be killed in a few

minutes by the use of chlorine. However, it doesn’t kill all of them for it

could withstand it up until a certain point before death. An example would

be Hepatitis A Virus, which could be killed in 16 minutes; E. Coli, killed in

less than 1 minute; and Giardia Parasite, killed in 45 minutes.

Based on the article of Ajora (2017), an indicator of fecal matter

would be the Total Coliform Bacteria. It is found on the variable however it

could be seen in the soil. There are different types of coliform bacteria

such as E.coli and Fecal coliform bacteria. Both are seen in the fecal

matter however the common bacteria would be E.coli. High levels of it will

rise due to the untreated sewage water, and poorly maintained septic

systems that have access to a mass of water.

Chemical Condition is another big indicator for it represents the

substances needed for a living thing in a specific area. It determines

whether the chemical composition of an area is able to sustain living

organisms. The subtopics that will be discussed are pH, Salinity, and

Dissolved Oxygen.

According to Ajora (2017), pH is a measure of how acidic or basic

the water is. It is meant as a negative log of hydrogen ions. It has a scale

that shows how acidic or basic a substance is. It ranges from 0-14 with 0

as most acidic and 14 as most basic. The number 7 is considered to be

neutral meaning it’s neither of the two.  

Similarly Fondriest (n.d), stressed the importance of pH to aquatic

life and water. The article stated that the pH level either too high or low will

kill any living organisms living within it. It also affects the way heavy metals

and chemicals dissolve in the area of water. It stated that even if the pH

had minor changes it could affect the environment and the organisms living

within it. It could stress the survival system of animals which could lead to

deaths and reduce reproduction. Another effect would be that if the pH

level is so high then it could increase the solubility of chemicals and heavy

metals which could turn toxic. 

According to DWER (2017), salinity is defined as the concentration

of salts in water or soil. The presence of salt in natural water is important to

aquatic life and high levels of salinity can alter the way water works.

However if acids are present then it could be harmful to many plants and

animals. The changes in land use, seasonal variation, climate changes can

affect the water and the amount of salt it contains. Salt comes into the

water by rain clouds or the landscape of the area. Rainclouds get the salt

which has evaporated from the ocean whereas the landscape could

contain rocks that has been weathered to contain salt.

According to Meadors (n.d), the effects of salinity are dependent if

the body of water is hyposaline or hypersaline. In terms of hypersalinity, it

could prove beneficial to marine life if the organisms can handle high

amounts of salt but if not then it could lead to reduction of fish growth.

Hyposalinity is the few amounts of salt in the water. It is beneficial for short

term development like combating parasites. However it could prove fatal to

other organisms like fish. Fish need to have an excessive amount of salt

for their kidneys but if in a hyposaline environment then its kidney system

could fail.

As claimed by James (2017), the factor has a unit of measurement.

Salt concentration is often measured in units of parts per million (ppm),

milligrams per liter (mg/L) , or percent. It could also be expressed in

practical salinity units (psu), a measure of conductivity at a constant

pressure and temperature. There are 3 methods which could help measure

it. The first would be the Conductivity Method. It states that the conductivity

of water is electrically proportional to its salt ions. It could be measured by

conductivity probe or meter and can be converted to measure salinity. The

second method would be Hydrometer Method. This mostly talks about the

water’s specific gravity will increase in proportion to the salt. Temperature

can affect the specific gravity and then converted into salinity. The

equipment used would be the hydrometer. Refractometer Method uses a

device called the refractometer. It estimates salinity by measuring the

degree the water can refract light.

Based on the article of Ajora (2017), dissolved oxygen is the amount

of oxygen dissolved in the water. It gets into the water by diffusion of air,

aeration and photosynthesis. It is a needed substance in water because of

aquatic life. The underwater plants get it first before the marine animals.

Some animals require high amounts of it in order to survive such as the

trout and stoneflies.

According to Fondriest Environmental Learning Center (n.d), the

level of dissolved oxygen varies due to the change of factors like

temperature. There are specific organisms living within the area that have

a certain limit to how much dissolved oxygen it could take. The minimal

amounts of oxygen being 1- 6 mg/L while the maximum being 4- 15 mg/L.

This limit shows that only specific animals could live within that area. It also

dictates that bacteria would also need dissolved oxygen just to decompose

organic material.

Physical Condition is an important factor to be considered for it

shows the state of the body of water from a general look on where it tests

the simple variables needed for a living organism to survive. According to

Ajora (2017), the condition is determined on the five senses which are

smell, touch, sight, sound, and taste. It has three subtopics like

Temperature, Color, and Total Suspended Solids.

According to Fondriest Environmental Inc (2014), water temperature

is property that expresses the hot or cold of a liquid. It measures the

thermal energy of a matter. Thermal energy is the kinetic energy of atoms

and molecules which is connected to temperature because as the atoms

are in an excited state, it becomes hotter whereas if in a cool, and neutral

state, it becomes colder. It’s measurement varies as it could be in Celsius,

Fahrenheit or Kelvin. It is an important factor because it could influence

other characteristics of water such as pH level, density, dissolved oxygen

and other dissolved gases, compound toxicity, metabolic rates and

photosynthesis production, conductivity and salinity, and oxidation

reduction potential. It also indicates what type of aquatic animal is present

to handle that sort of temperature. It states that the warmer the water gets

it could have less dissolved oxygen to be consumed whereas if cold water

it would have an increase of dissolved oxygen.

On the authority of Goff (2018), there are numerous tools to

measure the temperature of an object. The liquid expansion thermometer

is mostly a bulb or spring thermometer. It’s the most common tool for

measuring the heat. It works with alcohol or mercury which expands when

the temperature rises. Thermocouples are two metal leads of different

metals that when close to each other gives off an electrical charge. The

change in charge indicates the change of temperature. Resistance

Temperature Detectors are more stable compared to thermocouples. It

determines the change in electrical resistance caused by temperature

changes. Pyrometer measures the surface temperature of an object while

the Languir probe measures the plasma in the current. Finally the infrared
sensor measures it by looking at the heat signatures which determines the

temperature.

 In the opinion of  Trinex Internet Solutions Inc (2017), Color is a

water quality parameter that mostly focuses on the appearance of the

water. It is a natural material that dissolves into solution. It means that any

material could be present in the water but cannot be identified properly due

to its visual look. It could be the presence of metals, waste, and

microbiological matter that comes from outside sources. It mostly depends

on the look of the water which could help assume if its polluted or not. It

could be divided into two types of colors such as highly or lowly colored.

Highly colored cannot sustain aquatic life for long term but lowly colored

could sustain it. However the lowly colored would be harmful to humans

and can’t help identify which bacteria affects the area of water.

 According to Hazen (2017), he carried out an experiment for water

sewage treatment which created a color measurement index to quantify

the color of water. It is called the Hazen Color Scale. It is used by

comparing the water to a standard solution of diluted platinum cobalt. The

scale is from 0 (clean water) to 500 ( dark, polluted water). This is a very

helpful tool to determine the quality of water based on its color.

According to Fondriest Environmental, Inc.(2014), Total Suspended

Solids (TSS) are particles that are larger than 2 microns but if smaller then

they are dissolved. They are made up of inorganic material. They are

solids that are floating in the water from sediment or even from plants that

decays. They are a significant factor to consider when checking the water.

The high levels of solids then less chances of the water to be seen as

clear.  Total Suspended Solids is the main cause of turbidity. A way to

measure the TSS would be measuring their weight.It is a common method

to get the results. Its process would be to take  a water sample to be dried

then weighed. This is an accurate way to test the factor however it is a

time consuming and difficult one. It also affects the water clarity for if there

is a high suspended solids it could limit the sight of the aquatic organism

and show increased amounts of bacteria. Low suspended solids could help

the sight however it could show that the bacteria is lessened.

 Environmental Programs are movements that specialize in

rehabilitating or improving  the given natural environment. There are

various programs that focus on the water quality of a given body of water.

The organization, Environment Canada (2013), had created an

agreement called the “ 2012 Great Lakes Water Quality Agreement” to

which it serves as a treaty to analyze various areas of water and place

certain restrictions for its improvement. It was able to identify the area’s

problems using the Beneficial Use Impairments. This evaluation was used

to find out the causes of the declining water quality such as fish tumors,

stagnation of fish and wildlife population and many more. They also

established various lakewide management actions that keep the body of

water in a good condition. One of those actions is  to build a Lake

Ecosystem Objectives which would stand as a trademark to assess the

condition of the water. It also acquires the use of existing information as a

warning to show the state of the water. The team would need to research

and analyze which component could result in a greater threat within the

future.
 Likewise, Reganit (2019) has posted an article about the

rehabilitation of the Manila bay launched by the Department of

Environment and Natural Resources (DENR).  They launched the program

on January 27 and are expected to be finished by December. They have

gone to various phases and the first would be cleanup. They have pursued

and done different methods such as giving temporary sanitary facilities to

residents around the bay, cleaning up specific waterways, implementing

waste management, and relocating some residents. They were able to

educate the people to see the advantages of the program and have a fast

track record of completion. The current status would be that Pasig River is

currently being improved and the San Juan River is still infected with

different toxic waste. However employees believe that once Pasig River

and San Juan River is cleared then it will help the rehabilitation of Manila

Bay a lot faster. 

Whereas Gabbay (n.d), expresses that there have been movements

for rehabilitation of the Israel Rivers. The article states that a National

River Administration has been established to guide and improve the

restoration of the rivers. They have set standards for the water quality

which include the physical, chemical, and microbial parameters. An

example of a river would be the Yarkon River. They established the Yarkon

River Authority which has made actions for the river’s re-establishment.

They have removed hundreds of tons of trash,  have cleaned the water

banks and reinforced biking and hiking trails. The results of their actions is

shown by the return of flora and fauna, and has become an attraction site

of many picnics and historic sites.

Similarly, Komati Basin Water Authority or KOBWA (n.d), along with

DWS, NLM, MTA, and IUCMA, launched an awareness campaign program

called “River Dumping Awareness Campaign” that highlights the dangers

of river dumpings, issues of waste disposal, and protection of the natural

environment. They encouraged the people to find alternative ways to get

rid of their waste other than disposing of them on the river. An example

would be the area that surrounds Ingugwane River. They asked for

participants for the cleanup of the river. After the welcoming speech made

by the Councilor, the volunteers started cleaning up the area and removing

all trash within the vicinity. After the work, everyone was thanked and the

site has been seen as an example of how a water source should be kept

clean.
 Likewise Greater Kashimir (2017), stated that the M/S Montage

Traders Lal Mandi has launched an awareness program for lessening of

water pollution. They have shown various model that shows the reasons

for water pollution and how it could be controlled and managed. A

proprietor named Mehboob Beigh, stated that the campaign will show how

dangerous the waste is to the bodies of water. It would also present how it

could be minimize and recycled for other purposes. They also presented

equipment to students so they could know what to do in case of floods.

Similarly Batangas City Government (2020), they launched EBD

Magkatuwang Tayo program. The event aims to integrate the discipline

and good morals to the Batangas City Citizens. It would make people

prone to do the right thing, be it simple tasks like respecting elders and

throwing the garbage at the right place. It also has an effect towards

schools for the students are reminded to follow good discipline. It has put

the emphasis on good morals which influences the environment as well.

The program helps put into perspective the need to be disciplined on

where to properly put waste products. Overall it values the need of good

morals and discipline needed for the good of the city.

 Overall, the literature explains the importance of the variables in the

experiment. Each are important factors needed to measure the water

quality of the Calumpang River. The variables cover different areas of the

water which are biological, chemical, and physical aspects of it. The

environmental program is an integral part of the whole research for it is a

possible way to somehow rehabilitate the given river. All of the factors are

needed to fully explain why the river is the way it is.

Research Literature

According to Qureshimatva et. al (2015), their research, “Studies on

the Physico-Chemical Parameters and Correlation Coefficient of Sarkhej

Roza Lake, District Ahmedabad, Gujarat, India,'' was aimed to analyze the

status of the following bodies of water based on chemical characteristics.

They used factors such as temperature, ph, turbidity, total dissolved solids,

hardness, chlorides, phosphate, nitrates and biological oxygen demand. It

was done from September 2013 - 2014. It goes on to say the importance

of wetlands and the areas that will be studied by the researcher. It also

states the methods to acquire the sample and how they measured it. The

study concluded that the area was able to meet the necessary limits which

makes it usable for domestic and irrigation purposes.

 According to Hua (2017), his case study, “Potentially Land Used

Pattern Contributing Pollution Source Towards Water Quality:A Case

Study of UTM River,” showcases the state of the river and the causes of its

contamination. It describes how UTM River water quality has declined

because of various human activities on the land. It focused on the

dissolved oxygen, biological and chemical oxygen demand, suspended

solids and pH levels. The results showed that it’s in Class 1 and 2,

meaning that it is still considered safe water. However it could change due

to the human works such as agricultural and residential activity,

construction, and sedimentation. It goes on to say that humans should

respect the environment for if pollution is continued then it could lead to

environmental destruction. 

According to Kalshetty et.al (2014), their research, “Water Quality of

River Tungabhadra due to the Discharge of Industrial Effluent at Harihar,

District Davanagere, Karnataka State, India,” focuses on the quality of

water and the outside source affecting it. It goes on to say that the

importance of water is like how it’s the main factor of all activities of human

work. It also describes rain water as pure water even if it’s met with
dissolved gases. It explains how industrial work can change water

chemistry. Wastewater is produced and it comes with the addition of

organic matter, inorganic dissolved solids, fertilizer materials, suspended

solids, heavy metals, and microorganisms. It states that it contains

poisonous chemicals that could lead to the contamination of the water.

This study concludes that the standards are above the limited standard of

water and that an appropriate program is needed to fix this situation.

According to Adenji, O.O Okoh, A.I Okoh (2017), the study,

“Petroleum Hydrocarbon Profiles of Water and Sediment of Algoa Bay,

Eastern Cape, South Africa,” aimed to figure out the amount of Petroleum

Hydrocarbon in the water. It’s an organic matter in contaminants and can

be a good indicator of petroleum leftovers in masses of water. Petroleum

leftovers is essentially the oil that has been spilled out.  They gathered up

the contaminants and then analyzed it by their chemical properties. The

conclusion would be that the concentration of total petroleum hydrocarbon

has shown a slight rise of pollution which should be answered by a

meaningful law or act that could clear it up.

 According to Krestenitis et. al (2018), “Observational study of the

marine environment in the Northern Thermaikos Gulf,”  it limits itself only to

the Northern Thermaiko, inner and outer gulf, and aims to find its water

quality and the seabed. The samples were acquired by the placement of

two stations at the areas of the water treatment plant and at the city’s

seafront. The measurements for each sample included the physical,

biological, and chemical properties and seabed. The findings would be the

remarkable temperature and that high levels of phosphate and phosphorus

are present. In conclusion, the researchers were able to find the significant

difference between the inner and outer Gulf based on physical and

biochemical characteristics.

According to Hua, Kusin, & Praveena (2016), their investigation,

“Spatial Variation Assessment of River Water Quality Using Environmetric

Techniques,” focuses on the environmental techniques applied to

understand the size and diversity. The setting would limit to the Malacca

River and would use techniques such as hierarchical cluster analysis,

principal component analysis, and discriminant analysis. The first analysis

was used to group nine locations into two clusters based on their physico-

chemical and biological components whereas the second helped getting

the components such as pH, turbidity, temperature, and etc. The third

method was used to categorize the components among the groups. It was

discovered that residential areas and agricultural work are sources of

pollution in the first while sewage and industrial work are the other source

in the second. 

According to Hua, & Kusin (2015), “A Review in Analysis and

Detection of Water Quality Parameters in Malacca River. Case Study: A

Preliminary Finding,”  aims to analyze the water quality depending on its

chemical property. It was set up by having 9 stations in different areas, like

rivers and lakes, to collect selected variables. The results acquired depend

on the pH, dissolved oxygen, biological oxygen demand, and COD. The

conclusion would be that most of the results vary such that some factors

are considered to be pure while others are contaminated. It concluded that

the source of pollution would be factory activities, commercial work, and

livestock activity.

According to Bensig, Galapate, & Maglangit (2015), “ An

assessment of the organic pollution level of Buhisan, Bulacao and Lahug

rivers,Cebu, Philippines,” is testing the pollution level of the given rivers.

The parameters that were tested were pH, Biological oxygen demand, and

dissolved oxygen. It was compared with the DENR Administrative order

90-34 criteria for surface water. It was determined that the Bulacao River

was less polluted due to the fact that it had better solid waste management

and less amount of waste. It concluded that there is a need for

rehabilitation for the rivers from the community and the government.

The article of Dunca (2018) “Water Pollution and Water Quality

Assessment of Major Transboundary Rivers from Banat (Romania)”,

focuses on the water resources management and the need for laws that

protect the bodies of water. It goes further by providing an analysis of the

quality of water on the rivers from Banat. It also provides a method of

evaluation that is the Water Quality Index (WQI). It concluded that the

results presented show the water pollution and quality assessment from

the rivers and expressed the need for careful analysis of any given polluted

liquid source. That information could be used to find the appropriate

method of purifying the mass of water.

Hypothesis of the Study

 1. The biological, chemical, and physical condition of the Calumpang

River does not meet the DENR no. 2016-06 standard quality of

water.

Definition of Terms

The following terms are defined in this study:

 Biological Condition- it’s a condition that is an indicator of

watershed health such as aquatic life. In the study, the term refers to

one of the variables being studied.

 Chemical Condition- it’s the behavior of a substance that

undergoes chemical reaction. As used in the study, it refers to one of

the variables being studied.

 Physical Condition- it’s the condition or state of the body or bodily

functions that could be measured based on the five senses.

Operationally, it refers to being one of the variables being studied. 

CHAPTER II

RESEARCH METHODOLOGY

This chapter presents the method of preparation and

experimentation of the research.

Research Design

This paper is an Experimental Research. The researcher used a

standard procedure based on Standard Methods for the Examination of

Water and Wastewater. For this experiment, the researcher, together with

the staff of the Provincial Government and Natural Resources Office,

Batangas environment Laboratory, proceeded to two sampling stations

along the Calumpang River. The samples were analyzed to determine their

biological, chemical, and physical condition. Each condition has different

types which are the following variables: Fecal Matter, pH, Salinity,

Dissolved Oxygen, Temperature, Biological Oxygen Demand, Color, and

Total Suspended Solids. It was analyzed within a provincial testing center

and will follow their procedures. The experiment had three trials to test if

the results will stay consistent. After the results were recorded, it was

compared to a standard quality of water. Lastly, Standard deviation was

performed to get the results from the recorded analysis.

Apparatus and Materials

 Incubator set at 35.0 ºC + 0.5 ºC

  Water bath set at 44.5 ºC + 0.2 ºC

 Test tubes & test tube racks

 Durham tubes 

 Pipettes 

 Inoculating loop

 Beakers

 Dilution bottles 

 E. Coli (Positive control)

 YSI Model 3200 Instrument 

 Distilled Water

 Beaker , 100ml

 DO 300 Waterproof hand-held Dissolved Oxygen/Temperature

Meter 

 Distilled Water 

 pH meter with potentiometer, glass electrode and temperature

compensating device

 Beaker, 150 ml

 Stirrer / Stir bar

 Nessler tubes, matched, 50 –mL, tall form

 pH meter, for determining sample pH

 Filter and filter assembly

 Use a 0.45um-pore-diam cellulose membrane filter of 22 or

47 mm diam. Glass fiber filters also can be used. Rinse filters

before use and monitor filter blanks. Small- pore filters of 0.2

or 0.22 um or even ultrafiltration may be needed to remove

colloidal particles for certain samples such as Mn or Fe

oxides or other colloids. Use a glass, TFE#. Or stainless

steel assembly to hold the selected filters.

 Erlenmeyer or Iodine flask

 Buret, 25 mL

 Pipette, 1,10 ml

 Graduated Cylinder, 50 mL

 Analytical balance

 Incubation Bottles, 300 mL with glass toppers

 Incubation bottles, 250 or 300 mL with glass stoppers

 Air incubator or Water bath, thermostatically controlled at 20 1C.

Exclude all light to prevent the possibility of photosynthesis

production of D. O.

 Plastic Container, 10L or more

 Air Pump, twin Outlet

  Pipette,  5, 10 mL

 Beakers and Volumetric flasks

 Graduated Cylinder , 100 mL

 Buret , 25 mL

 Apparatus listed in 2540B.2and C.2 is required, except for

evaporating dishes, steam bath and 180°C drying oven. In addition:

Aluminum weighing dishes

Chemical/Reagents

 Lauryl Sulfate Broth 

o For triple strength, weigh 106.8 g Lauryl Sulfate Broth.

o Dissolve in 1L with distilled water and stir.

o Dispense 10ml broth in test tubes fitter with Durham Tubes.

o Autoclave for 15 minutes at 121ºC and 15 psi.

o Cool at room temperature before refrigerating.

o Ph after sterilization should be 6.8 + 0.2.

 Dilution Water (0.1% peptone H2O) 

 o Dissolve 1.0 g of peptone per liter of distilled water. Dispense

100ml of  solution to 125ml autoclavable bottles. 

o Autoclave for 15 psi, at 121 ºC 15 minutes. 

 pH 4.0 Buffer

 pH 7.0 Buffer

 pH 10.0 Buffer

 KH2PO4 solution as quality control sample

 Fry KH2PO4 in desiccator and weigh 54.436 g in a clean and

dry beaker. Transfer into 1-L volumetric flask. As 180.23 ml

1.0 NaOH. Make a solution up to volume with distilled water.

Dilute 5 ml aliquot to 100 mL distilled water. The solution will

produce 239 mg/ L total dissolved solids, 3036 umhos/cm and

pH of 6.87.

 Stock Standard (500PCU)

 Dissolve 1.246 g potassium chloroplatinate, K2PtCl6 and 100

g crystallized cobaltous  chloride, CoCl2.6H2) in water with

100 mL concentrated HCI. Dilute to 1000 mL. This stock

standard has a color of 500 color units.

 Working Standards
  Prepare having CU of 5,10,15,20,25,30,40,50 and 100 by

diluting 1.0, 2.0, 3.0, 4.0, 6.0, 8.0, 10.0, 20.0 stock color

standard with distilled water to 100 mL volumetric flask.

Protect standard against evaporation and contamination when

not in use. Keep in the dark when not in use, and keep only for

1 month.

 Alkali-iodide-azide reagent

 Dissolve 5g NaOH ( or 700 Potassium Hydroxide, KOH) and

135 Nal (or 150g Potassium iodine, Kl), in distilled water dilute

to 10000 mL and cool.

 Slowly, with stirring, add solution of 10g NaN3 dissolve in 40

mL of water. Chemically equivalent potassium salts may be

used interchangeably. Diluted and acidified solutions  must not

give color with starch indicator.

 Store in dark blue with a rubber stopper.

 Manganese Sulfate Solution

 Dissolve 364g Manganese Sulfate Monohydrate,

 Filter and dilute to 1 liter. No more than trace of I2 should be

liberated when solution is added to acidity Kl solution.

 Starch Indicator Solution

  Dissolve 2g soluble starch and 0.20g salicylic acid, as a

preservative in 100 mL hot distilled water.

 Sulfuric Acid, concentrated

 Standards Sodium Thiosulfate Titrant

 Dissolve 6.205 g Sodium Thiosulfate Pentahydrate, in freshly

boiled and cooled distilled water. Add 1.5 mL 6N Sodium

Hydroxide, NaOH or 0.4 g solid NaOH and dilute to 10000 mL.

*(exactly 0.025 N thiosulfate is exactly 0.025 N thiosulfate is

equivalent to 0.200 mg D.) per milliliter) Standardized with

0.0021 M potassium bi-iodate solution.

 Standard Potassium bi-iodate solution, 0.00021M

 Dissolve 812.40 mg KH in distilled water dilute to 1000 mL.

 Standardization of Sodium Thiosulfate (0.025N)

 Dissolve 2g kl., Free from iodate , in a 250  m l Erlenmeyer

flask with 100 to 150 ml distilled water. 

 Add 1 mL 6N H2SO4 or a few drops of concentrated H2SO4

and 20.00 mL of standard bi-iodate solution.

 Dilute to 200mL and titrate liberated iodine with thiosulfate

titrant, adding starch toward the end of titration. When a pale

straw color is reached.

 Dilution Water – used distilled water

 Phosphate Buffer Solution – pH 7.2

 Dissolve 8.5 g Potassium DihydrogenPhosphate, KH2PO4,

21.75 g Dipotassium Hydrogen Phosphate, K2HPO4, 33.40 g

Disodium Hydrogen Phosphate heptahydrate,

Na2HPO47H2O, and 1.70g Ammonium Chloride, NH4CI in

about 500 mL distilled H2O and dilute to 1000 mL. discard

reagent (or any of the following reagents) if there is any sign of

biological growth in the stock bottle.

 Magnesium Sulfate Solution

 Dissolve 22.50 g Magnesium Sulfate Hexahydrate, in distilled

water and dilute to 1000mL.

 Calcium Chloride Solution

 Dissolve 27.50g Calcium Chloride, CaCl2, in distilled water

and dilute to 1000mL.

 Ferric Chloride Solution

 Dissolve 0.25g Ferric Chloride Hexahydrate, FeCl3-6H2O in

distilled water and dilute to 1000mL.

 Acid and Alkali Solutions, 1N

 For neutralization of caustic and acidic waste samples.

  Acid – slowly and while stirring, add 28mL conc. Sulfuric Acid,

H2SO4 to distilled water and dilute to 1000 mL.

 Alkali – dissolve 40g Sodium Hydroxide, naOH, in distilled

water, Dilute to 1000mL.

 Sodium Sulfite  Solution

 Dissolve 1.575g Sodium Sulfite, NaSO3, in 1000 mL distilled

water. This solution is not stable; prepare fresh daily.

 Glucose-Glutamic Acid Solution

 Dry reagent grade Glucose and Glutamic Acid at 1036C for 1

hour. Dissolve 0.15g glucose and 0.15g glutamic acid with

distilled water and dilute to 1L. prepare fresh daily.

 Polyseed Solution

 Transfer content of the polyseed capsule to 500 mL of distilled

water. Aerate  with an air pump for 1 hr.

 Ammonium Chloride Solution

 Dissolve 1.15g NH4CL in about 500mL distilled water, adjust

pH to 7.2 with NaOH solution and dilute to 1L. Solution

contains 0.3 mg N/ Ml/

Procedures
Sampling Steps:

1.   Put on protective gloves and wading boots.

2.  Wade into the water to the center of the river channel where the water

is deepest and current has the greatest velocity. Face upstream and wait

until the plume of sediment has been carried away or has settled. 

3.  Rinse the container at least three times with the river water, throwing

the used water downstream of the sampling location.

4.  Lower the sample container into the water face down. Hold it with one

hand on each side to a depth at least 4 inches below the surface or

halfway to the bottom of the stream. . Do not touch the inner part of the

container. If the stream is very shallow, lower to a depth just above the

stream bed but do not touch or disturb the stream bed with the container.

5.   Slowly lift the container towards the flow. Fill it to about 4/5 full. Enough

space should be left to allow for addition of preservative, if necessary, and

to allow for mixing the sample.

6.  Cap or cover the container and bring the sample to the working area for

the succeeding steps. 

Fecal Coliform

1. Preservation of Sample  

a.  For E. coli analyses, the holding time from collection to analysis is

30h.

b.  Keep samples at 8<ºC during transport to the laboratory. 

   2. Presumptive Phase

a. Arrange fermentation tubes containing Lauryl Sulfate Broth in rows

of five each in a test tube rack per 10ml dilution. 

b. Shake sample and dilutions vigorously about 25 times.

c. Inoculate each tube in a set of five with 10ml of corresponding

dilution and mix test portions in the medium by gentle agitation. 

d. Incubate inoculated tubes at 35 + 0.5 ºC. after 24 + 2h swirl each

tube gently and examine for growth, gas and acidic reaction (shades

of yellow color) and if no gas acidic reaction is evident, reincubate

and reexamine at the end of 48 + 3h. Record presence or absence

of growth, gas and acid production.

e. Production of an acidic reaction or gas in the tubes within 48 + 3h

constitutes a positive presumptive reaction.  Submit tubes with a

positive presumptive reaction to the confirmed phase. The absence

 of acidic reaction or gas formation at the end of 48 + 3h of incubation

constitutes a negative test.  

3. Confirmed Phase

a. Submit all presumptive tubes or bottles showing growth, any amount

of gas or acidic reaction within 24 + 2h of incubation to the confirmed

phase. Submit additional tubes showing active fermentation or acidic

reaction at the end of a 48 + 3h incubation period to the confirmed

phase. 

b. Gently shake or rotate presumptive tube showing gas or acidic

growth to resuspend the organisms. 

c. With a sterile loop 3.0 to 3.5 mm in diameter, transfer one or more

loopfuls of culture to a fermentation tube containing Brilliant Green

Lactose Bile Broth. Repeat for all other positive presumptive tubes.

d. Incubate the inoculated brilliant green lactose bile broth tube at 35+

0.5 ºC.

e. Formation of gas in any amount in the inverted vial of the brilliant

green lactose bile broth fermentation tube at anytime within 48 + 3h

constitutes a positive conformed phase.

 f. Calculate the MPN value from the number of positive brilliant green

lactose bile tubes as described in 5.5. 

Alternative procedure: Use this alternative only for polluted water or

wastewater known to procedure positive results consistently. If all

presumptive tubes are positive in two or more consecutive dilutions

within 24h, submit to the confirmed phase only the tubes of higher

dilution in which all tubes are positive and any positive tubes in still

highest dilutions. Submit to the confirmed phase all the tubes in

which gas or acidic growth is produced only after 48 hours. 

4. Completed Phase

a. Use the completed test on at least 10% of positive conformed

tubes. 

b. noculate simultaneously into Brilliant Green Lactose Bile Broth for

total coliforms and EC broth for fecal coliforms.

c. Consider positive EC broth at elevated temperature (44.5 ºC) results

as a positive completed test response. 

5.  Fecal Coliforms

 a. Submit all presumptive fermentation tubes showing any amount of

gas, growth or acidity within 48h of incubation to the fecal coliform

test.   

b. Gently shake or rotate presumptive fermentation tubes showing gas,

growth or acidity.   

c. Using a sterile 3.5 mm-diameter loop, transfer growth from each

presumptive fermentation tube to EC broth. 

d. Incubate inoculated EC broth tubes in a water bath at 44.5 + 0.2 ºC

for 24 + 2h. Place all EC tubes in water bath within 30 minutes after

inoculation. Maintain a sufficient water depth in water bath incubator

to immerse tubes to upper level of the medium.  

e. Gas production with growth in an EX broth culture within 24+ 2h or

less is considered a positive fecal coliform reaction. Failure to

procedure gas (with little or no growth) constitutes a negative

reaction.   

f. Calculate MPN from the number of positive EC broth tubes as

described in 5.5. 

Salinity
1. Make sure that the instrument is set up to read the units you wish to

measure (in conductance, conductivity, resistivity, salinity, etc.)

2. To change the units, from the Setup Disp Screen, press the [NEXT]

soft key to select the field you want to change. Press [ENTER]

repeatedly to change the field until the formation you want is

displayed.

3. Verify that the 3200 is properly set up to use the current cell by

measuring calibrating with standard conductivity solution.

4. Immerse the cell in the solution to be measured. If using a YSI 3200

series cell and correcting for temperature, also place the

temperature probe in the solution. 

5. Gently tap the cell to remove any air bubbles. 

6. Dip the cell in the solution 2 or 3 times to ensure proper wetting. The

cell electrodes must be submerged and the electrode chamber must

not contain any trapped air. If using a flow through or fill cell, be

certain it is completely full.

7. Allow time for the temperature to stabilize.

8. Read the display, send the reading to a computer or serial printer, or

store the reading in memory.

 9. Rinse the cell (and the temperature probe, if used) with distilled H2O.

Immerse the cell in distilled water.  

Dissolved Oxygen

1. Dissolved Oxygen Calibration in % Saturation 

a. To calibrate 100% Saturation, press MODE key to select %

saturation mode. 

b. Rinse the probe well with distilled water. Wipe the end of the probe

dry. 

c. Hold the probe in the air with the sensor facing down and press CAL

key to calibrate the meter. 

d. The LCD will show a “CAL” mode. The primary display will show the

current value of measurement and the secondary display will show

“100.00” to which the meter is going to be calibrated. Wait for the re

adding to stabilize.

e. Press ENTER key to confirm the calibration. The meter automatically

calibrates to 100% air saturation and returns to measurement mode.

2. Taking DO Readings 

a. Dip the probe into the sample. Stir the solution gently using the

probe to homogenize the sample. 

 b. Note the reading on the display.

Temperature

1. Sample analysis

a. Place 100ml beaker and dip electrode alongside calibrated

thermometer to check correctness of reading

b. Stir gently to ensure homogeneity, immerse electrode and

thermometer in sampling long enough to permit complete equilibrium

c. Record temperature reading

d. Clean electrode with distilled water and blot dry.

Color

1. Estimate of intact sample

a. Check sample pH.  If outside the range of 4 to 10, preferably

adjust sample to pH7 and  note the adjustment. If true color is

to be measured , wash membrane filter and filter assembly by

passing at least 50 mL water through filter. Filter about 25 mL

sample and discard filtrate. Filter a further portion of about 50

mL through the same filter and retain for analysis.

2. Observe sample color by filling a matched nessler tube to the

50ml mark with sample and compare it with standards.

 3. Look vertically downward through tubes toward a white or

specular surface placed at such an angle that light is reflected

upward through the columns of liquid.

4. If turbidity is present and has not been removed, report as

“apparent color”. If the color exceeds 100  units, dilute sample in

known proportions until the color is within the range of the

standards.

pH

1. Calibrate pH meter using pH buffers 4.0,7.0, and 10.0.

a. Choose either 4.01 and 7.00 or 7.00 and 10.01 buffers,

whichever will bracket your expected sample range.

b. Rinse electrode and place into the 7.00 buffer.

c. Press MODE key until calibrate is displayed

d. Press YES key to use the sequence

e. When READY is displayed, press YES key. Then P2 is being

displayed

f. Rinse the electrode and place into the second buffer

g. When READY is displayed, press the YES key

h. After the second buffer value has been entered, the electrode

slope will be displayed

 i. The meter will advance to the measure mode.

Biological Oxygen Demand

1. Preparation of Dilution Water

a. Determine volume of distilled water to be used and place in suitable

container (6300 mL of distilled water of 1 sample, just add a1800 mL

for every 1 sample added.)

b. Add 1 mL each of Phosphate Buffer, MgSO4, CaCl2 and FeCL3

solutions per 1000ML of water.

c. Before use, bring dilution water temperature to 10C

d. Saturate with dissolved oxygen by aerating with an air pump for 1 hr.

2.   Dilution Water Storage

a. Source water maybe stores before use as long as the prepared

dilution water meets quality control criteria in the dilution water

blank.

b. Preferably do not store prepared dilution water for more than 24h

after adding nutrients, minerals, and buffer unless dilution water

blanks consistently meet quality control limits.

3. Dilution Water Blank

a. Use a dilution water blank as a rough check on quality of unseeded

dilution water and cleanliness of incubation bottles.

b. Siphone dilution water into BOD bottle to full. Do not add seed

solution. Prepare two sets of dilution  water blank.  Water seal one

bottle and incubate for 5 days at 20 1C. determine the initial DO of

the other set. The DO uptake should not be more than 0.20 mg/L.

4. Glucose –Glumatic Acid – Quality Control Sample Check

a. Siphon dilution water into BOD bottle to one-fourth full. Make 2 sets

of samples with 3 dilution.

b. Add 6 mL of polyseed solution to each bottle. (always check the

volume of polyseed to be used in the information sheet provided in

the InterLab Polyseed Application Procedure)

c. Fill with dilution water.

d. Water seal the bottle and incubate one set for 5 days at 20 1C

determine the initial DO of the other set.

e. For the 300 mg/L mixed primary standard, the average 5-day BOD

would be 198 mg/L with a standard  deviation of 30.5 mg/L.

5. Seed Control Factor (SCF)

a. It is necessary to be present  in each BOD bottle a population of

microorganisms  capable of oxidizing the biodegradable organic

matter in the sample.

b. PG-ENRO BEL identified the street canal as source of  seeding as a

replacement of SCF(polyseeds). The performance of the seed in

BOD test has been tested and passed the criteria of having a DO

uptake attributable to the seed of 0.6-1mg/L. thus it is used as Seed

Control Factor.

c. Siphon dilution water into BOD bottle to one –fourth full. Make 2 sets

of 3 dilutions.

d. Fill with dilution water.

e. Water seal the bottle, incubate one set for 5 days of 20 1C and

determine the initial DO of the other set.

6. Sample Pre-treatment

a. Check pH of the sample before testing. If it is not between 6.0 and

8.0 adjust sample temperature to 20 +-3C, then adjust pH to 7.0-

7.20 using a solution of sulfuric acid or sodium hydroxide of such

 strength that the quality of reagent does not dilute the sample by

more than 0.5%. Always seed sampled that have been pH adjusted.

b. If the sample has been chlorinated but no detectable chlorine

residual is present, seed the dilution water. If the residual chlorine is

present, dechlorinate the sample and seed the dilution water.

Destroy chlorine residual by adding Na2SO3 solution. Determine the

required volume on a 100-to 1000mL portion of neutralized sample

by adding 10mL of 1+1 acetic acid or 1-50 sulfuric acid, 10mL

potassium iodine (kl) solution to the starch –iodine endpoint for

residual. Add the neutralized sample to the relative volume of

na2SO3 solution, mix and  after 10 to 20 min check sample for

residual chlorine.

c.  Samples containing more than 9 mg DO/L at 20 C must be

prevented to the loss of oxygen during  incubation by bringing the

sample to about 20+3C in partially filled bottles while agitating by

vigorous shaking  or by aerating with clean filtered compressed air. 

7. Sample Preparation

a. Siphon dilution water into a BOD bottle to one-fourth full. Make at

least three dilutions of prepared samples to produce a residual DO

 at least 1.0mg/L and a DO uptake of at least 2.0mg/L after 5-d

incubation. 

b. Add 4mL of polyseed solution to each bottle. (always check the

volume of polyseed to be used in the information sheet provided in

the InterLab Polyseed Application procedure.

c. Refer at the table below for dilutions to be prepared. Compute the

volume of the sample based on the percent dilutions desired.

d. If DO is zero and appearance of sample showed to be highly

polluted, prepare  5-10% dilutions of the sample.

e. Using a volumetric pipet, add the desired volume of sample to each

BOD bottles.

f. When a bottle contains  more than 67% of the sample after dilution.

Add nutrient, mineral and buffer solutions directly to the diluted

sample at a rate of 1mL/L (0.30mL/300mL bottle).

g. Add dilution water to full. Water seals the sample.

8. Sample Incubation

a. Incubate one set for 5 days at 20C at 1C. Exclude light to avoid

growth of algae in the bottles during incubation.

9. Determination of Initial DO
 a. After 5 days of incubation , follow the steps on the determination of

initial DO to determine the final DO.

b. Quality  Control Checks

c. Minimum residual DO and minimum DO depletion; only bottles,

including seed controls, giving a minimum DO depletion of 2.0mg/L

and a residual DO of at least 1.0mg/L after 5d of incubation are

considered to produce valid data.

d. Glucose-glutamic acid check: the resulting average BOD for three

bottles must fall into the range of 198 +30.5mg/L.

e. Dilution water quality check: the DO uptake in 5d must not be more

than 0.20mg/L and preferably not more than 0.10mg/L.

f. Seed control: The DO uptake attributable to the seed added to each

bottle generally should be between 0.6 and 1.0 mg/L.

Total Suspended Solids

1.  Preparation of glass-fiber disk: If pre-prepared glass fiber filter disks

are used, eliminate this step. Insert disk with wrinkled side up in filtration

apparatus. Apply vacuum and wash disk with three successive 20-mL

portions of reagent-grade water. Continue suction to remove all traces of

water, turn vacuum off and discard washings removed filter from filtration
apparatus and transfer to an inert aluminum weighing dish. If a Gooch

crucible is used, remove crucible and filter combination. Dry in an oven at

103 to 105°C for 1 h. If volatile solids are to be measured, ignite at 550°C

for 15 min in a muffle furnace. Cool in desiccators to balance temperature

and weight. Repeat cycle drying or igniting, cooling, desiccating and

weighing until a constant weight is obtained or until weight change is less

than 4% of the previous weighing or 0.5, whichever is less. Store in

desiccators until needed.

2.  Selection of filter and sample size: Choose sample volume to yield

between 2.5 and 200 mg dried residue. If volume filtered fails to meet

minimum yield, increase sample volume up to 1L. if complete filtration

takes more than 10 min, increase filter diameter or decrease sample

volume. 

Recording of Data

1. Acquire Data after the analysis

2. Input statistical treatment on the data

3. List it down.

Statistical Treatment of Data

 The data that were recorded resulting from the experiment were

treated using the following:

1. The data will be recorded and differentiated in tables, descriptions,

and pictures.

2. The experimentation and analysis of the water will be observed and

recorded.

3. The researcher compared the results to the standard water quality of

the DENR no. 2016-06.

4. Mean, Standard deviation, Standard Error Mean,and T-test were

used to determine and identify the difference of the results to the

standard.
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