LAB 7 – Water Quality: Biological Characteristics of Water
Pre-laboratory Questions – Answers can be found in the Lab Instructions Background, or in your text.
1. Why do we test water for ph, DO, BOD, turbidity and coliform?
2. Discuss human activities that could affect the quality of the natural surface water at your test site. Specifically address
temperature, dissolved oxygen, coliform bacteria, BOD and turbidity.
3. Briefly describe how each of the following parameters interacts with at least one other factor/parameter (for example, how
does temperature affect DO?).
a. Temperature
b. Dissolved Oxygen
c. Coliform Count
d. Biochemical Oxygen Demand
e. Total Suspended Solids or Turbidity
Data Table: Water Quality: Biological Data and Site Photos
Test
Site
Site Name
or
GPS location
Elev.
(ft)
Temp
(°C)
Dissolved
Oxygen
(ppm)
DO
%
Saturation
BOD
(ppm)
pH
1
Coliform
Bacteria
(positive
or
negative
)
Water
Vol. for
Turbidity
Turbidity
(NTU)
Notes on Site 1 - insert a picture of the body of water you sampled
2
Notes on Site 2 - insert a picture of the body of water you sampled
Observation Questions
4.
Describe your sampling location. Was the water still or running? If it was moving, how fast (generally)? Was the water clear or
cloudy? Was there any algal growth? What about the surrounding land? Was it urban, suburban, or rural? Grassy or forested?
5.
How could your sampling location affect the quality of the water in your sample?
Discussion Questions
6.
RESULTS - Using your data above, How healthy are the water bodies you tested? (Hint, discuss your data results and use them
to come to a conclusion).
7.
Do all “healthy” bodies of water have the same biological data? Explain.
8.
Residents in a riverfront community have complained about a recent decrease in water clarity. Local ire has fallen on a local
pig farm that has recently expanded its operations upstream of the town, but the pig farm claims that it is taking actions to
protect the waterway and is pointing to the new shopping center across the river as a source of increased sedimentation. What
water quality tests could you perform to determine the source of the increased turbidity? How would you expect the samples
to differ if the pig farm is responsible?
9.
If you could do a biological water quality test on surface water anywhere in the world, where would you test and why?
Quality of Surface Waters:
Biological
Carolina Distance Learning
Investigation Manual
©2015,Carolina Biological Supply Company
Table of Contents
Overview ....................................................................................................... 4
Outcomes ..................................................................................................... 4
Time Requirements ...................................................................................... 4
Background .................................................................................................. 5
Materials ........................................................................................................ 9
Safety ........................................................................................................... 10
Preparation ................................................................................................. 10
Activity 1: Temperature Measurement .................................................. 11
Activity 2: Sample Collection .................................................................. 12
Activity 3: Dissolved Oxygen and Biological Oxygen Demand Test 12
Activity 4: Turbidity Test ............................................................................. 15
Activity 5: Coliform Bacteria Test ............................................................ 15
Disposal and Cleanup .............................................................................. 16
Data Table: Biological Water Quality .................................................... 18
©2015,Carolina Biological Supply Company
Overview
Students will collect water samples from local waterways and test them for biological
factors that governments and scientists use to monitor the health of natural waterways.
Outcomes
•
Practice proper water sample collection techniques.
•
Conduct chemical and biological analyses of field water samples.
•
Compare water test results to environmental standards.
Time Requirements
This lab requires the collection of water samples at two sites, travel time is in addition to
any times listed below.
Preparation.............................................................................................40 minutes
Activity 1: Temperature Measurement ...............................................10 minutes
Activity 2: Sample Collection ...............................................................10 minutes
Activity 3: Dissolved Oxygen and BOD Test .......................................20 minutes + 5 days
Activity 4: Turbidity Test .........................................................................10 minutes
Activity 5: Coliform Test .........................................................................10 minutes + 48
hours
©2015,Carolina Biological Supply Company
Background
Water is one of the most common compounds on earth, and one of the most
precious. Water covers approximately 75% of the earth’s surface, but only 2.5% of this
is fresh water, and only a small fraction of that 2.5% is accessible for use and
consumption. We depend on a clean water supply for drinking, cleaning, sanitation,
recreation, and manufacturing. Public health officials and biologists have developed
tests that can be used to monitor the quality of natural waters and to locate sources
(and potential sources) of water pollution. In this activity, you will collect samples from
local waterways and test and evaluate water quality on the basis of several chemical
and physical characteristics used by scientists to analyze natural waters.
Nitrogen and Phosphorous
Water is an important agent for dissolving and transporting many nutrients (essential
compounds for life) through the environment. The growth of primary producers (and,
by extension, all organisms in the food chain) in any environment is limited by the
scarcity of a particular resource. That resource can be physical, such as light or space,
or it can be a nutrient. In terrestrial and freshwater environments, nitrogen or
phosphorous often is limiting. Most fertilizers contain nitrogen and phosphorus as watersoluble nitrates (NO3–) and phosphates (PO43–). Rain washes these water-soluble
nitrates and phosphates from cultivated land into surface waters where they promote
the rapid growth of plants and algae, which speeds up the rate of eutrophication.
Eutrophication is a natural, usually gradual process that increases sediment and
nutrient availability in water bodies. Human activities lead to cultural eutrophication.
Fertilizer runoff from lawns and farms, inadequately treated municipal wastewater,
and poorly maintained septic systems all contribute to cultural eutrophication. In
agricultural areas with dense animal populations, animal waste adds nitrogen and
phosphorus to surface waters. Heavy rain can cause a sudden influx of nutrients,
which can lead to rapid and uncontrolled growth in aquatic algae and plant
populations. When these organisms die, bacterial-mediated decay depletes water
oxygen levels, kills sensitive aquatic organisms, and contributes to sediment buildup. If
nitrogen is the limiting nutrient, such as in most brackish or marine waters, nitrogen
levels ≥ 0.3 parts per million (ppm) can accelerate natural eutrophication.
Phosphorous is usually the limiting nutrient in freshwaters. Phosphorus levels ≥ 0.1 ppm
can accelerate natural eutrophication in freshwaters.
Temperature
Temperature affects many chemical and biological processes. It modifies the solubility
of oxygen in water, the rate of photosynthesis in aquatic plants, the metabolic rate of
aquatic organisms, and the susceptibility of aquatic species to toxins and disease.
Water temperature varies naturally with season and depth, but human activity also
can affect temperature in significant ways. For example, heated water from surfaces
such as sunlit parking lots, roads, sidewalks, and from industrial processes that use
water as a coolant can flow into nearby creeks, streams, or rivers. This thermal
pollution can destroy some types of aquatic life. Water loses heat very slowly, so
thermally polluted water should be given significant dilution and time to cool before
discharge into natural surface waters. Heated water also facilitates the rapid growth
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of some aquatic plants and algae, so thermal pollution can accelerate the rate of
cultural eutrophication.
Dissolved Oxygen
Dissolved oxygen (DO) is essential for biotic diversity in a body of water. The amount of
oxygen in water is usually expressed in ppm. Much of the DO in water comes from the
atmosphere. Atmospheric oxygen dissolves via diffusion into the water at the surface.
Wind and waves stir the water, which dilutes the DO concentration at the surface by
mixing, and subsequently increases the amount of oxygen that can dissolve at the
surface. Another important source of oxygen in water is photosynthesis performed by
aquatic plants and algae. Some of these aquatic photosynthetic organisms are
rooted or attached to the streambed, whereas others are planktonic (live in the water
column). The amount of photosynthetically generated DO in the water typically peaks
in the afternoon when the sun reaches zenith and the most light penetrates the water.
Turbidity, or murkiness resulting from a large number of particles in the water, can
lower DO levels by reducing photosynthetic rates due to limited light penetration.
Many factors influence the amount of DO water can hold. Water with lower
temperatures can contain higher oxygen concentrations. Freshwater can hold more
oxygen than saltwater. Another factor is altitude and atmospheric pressure; higher gas
pressures result in higher gas concentrations dissolving into a liquid. Atmospheric
pressure is greatest at sea level, so more oxygen dissolves into water at sea level than
at higher elevations.
The amount of DO in a body of water can vary greatly due to all these factors. After a
DO measurement is taken from a water sample, a percent saturation value can be
calculated by comparing the amount of oxygen the water contains at that moment
with the maximum amount of oxygen that it could contain at that temperature.
Respiring organisms such as animals and bacteria significantly reduce DO levels.
Bacteria in animal waste are a major concern in waterways that receive runoff from
fields and pastures where livestock are kept. When oxygen becomes depleted in the
water, it is said to be anoxic. Anoxic conditions cause marine organisms to suffocate.
Coliform Bacteria
Coliform bacteria populate the intestinal tract of warm-blooded animals, and disperse
throughout the environment along with fecal material. When an animal defecates,
large numbers of coliform bacteria (called fecal coliforms) are present in the feces.
Coliforms also can be released to the environment from a decomposing body. The
presence of large numbers of coliforms in a water sample is a good indication that
animal or human waste has polluted the water. Most coliforms do not cause disease,
except for a few Escherichia coli strains. However, many pathogenic bacteria and
parasites are spread through feces (e.g., those causing typhoid fever, hepatitis,
gastroenteritis, and dysentery), and the presence of coliforms indicates that harmful
pathogens might be present. Determining the presence of coliforms in drinking water
and in swimming areas is a very important tool for protecting public health.
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This is accomplished by collecting a water sample, and inoculating some of the
sample on nutrient rich medium that supports coliform growth. After several days, the
culture will show bacterial growth. This is the first step in determining the risk of animal
or human waste contamination of a body of water. Additional tests can be performed
to distinguish between total coliforms, which can have a number of sources, and fecal
coliforms, which originate in animal and human intestines. There are potential risks
associated with culturing pathogenic bacteria. Therefore, this activity will only
examine total coliforms.
Total Suspended Solids
Total suspended solids (TSS) is a term used to describe and measure the material left
behind after a water sample has been filtered and evaporated. Dissolved materials
and nutrients in natural water come from a variety of sources. Large concentrations of
dissolved solids affect cellular nutrient transport, alter the taste of water, and interfere
with industrial processes by fouling pipes and filters. Calcium and magnesium are two
minerals commonly dissolved in water. A water-insoluble carbonate forms when
aqueous calcium or magnesium hydrogen carbonate is heated. Industrial boilers
reach high temperatures and generate solid deposits of magnesium carbonate and
calcium carbonate, commonly called scale. These water-insoluble deposits reduce
the efficiency of machines and are expensive to remove. TSS measurement requires
the use of an analytical balance, which is not practical in the current setting due to
cost.
Turbidity
The turbidity test measures water clarity. Fine particles such as eroded soil, and
microscopic organisms such as phytoplankton, contribute to water turbidity. Turbid
waters clog fish gills, hinder disinfection processes, and carry entrapped pollutants
downstream. Turbidity also blocks sunlight, which prevents photosynthesis below the
surface and increases surface water temperature. Turbidity is measured in
Nephelometric Turbidity Units (NTU). In this exercise, a modified turbidity tube will be
used. A water sample is drained from the tube slowly, and the black and white
pattern of a Secchi disk at the bottom of the tube becomes visible. At this point when
the disk becomes visible, the sample volume is recorded and translated into NTU. If
the pattern is visible when the tube is full, turbidity will be estimated using a saturation
chart.
Biochemical Oxygen Demand
Aerobic bacteria that decompose organic material require oxygen. In general, the
more organic material there is in the water the oxygen demand will increase. When
biochemical oxygen demand (BOD), sometimes called biological oxygen demand,
surpasses the amount of oxygen that can be added to the water, the water becomes
anoxic. High BOD coupled with low DO input causes water quality deterioration,
which can result in the death of many fish and animals.
An increase in nitrates and phosphates in a body of water often increases the growth
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rate of plants and algae. These aquatic organisms subsequently die and are
decomposed by aerobic bacteria, which consume oxygen during the process.
Another cause of increased BOD is a consequence of fertilizer runoff. An influx of
fertilizers and sudden growth of aquatic producers may produce a brief spike in
photosynthetically generated DO levels, but BOD rises as the aquatic producers begin
to die.
pH
Water contains hydrogen ions (H+) and hydroxyl ions (OH–). The concentration of H+
ions determines the pH. If the concentration of H+ ions is higher than that of OH– ions,
the pH is less than 7.0 and is considered as acidic. Thus the greater the concentration
of H+ ions, the lower the pH will be. If the concentration of H+ ions is lower than that of
OH– ions, the pH is greater than 7.0 and is considered as alkaline (or basic). The pH
scale is from 0 to 14. The pH scale is logarithmic; as the pH increases by one whole
number, the H+ ion concentration decreases 10 times. For example, a pH change from
5 to 6 indicates a tenfold decrease in H+ concentration, and a pH change from 4 to 6
indicates a hundredfold decrease in H+ concentration.
Water rarely exists at the neutral pH of 7. Saltwater typically is slightly basic, with pH of
8 or higher. Freshwater usually has pH of 6.5, but this varies greatly between bodies of
water. Organic materials can affect the pH of water. Fallen leaves often contain
tannic acid, which stains the water brown and reduces pH. Some bacteria create
acidic waste products that lower the pH of water. Acid rain has a widespread effect
on natural waterways in the eastern United States. Oxides of nitrogen and sulfur are
released into the atmosphere by burning fossil fuels. They mix with atmospheric water
and form nitric and sulfuric acids. These acids are mixed with rainwater and delivered
to natural waterways and soils. Normal rain has pH of 5.6 (due to dissolved carbon
dioxide from the atmosphere forming carbonic acid). Rain is considered to be acid if
the pH is lower than 5.6.
Many freshwater organisms have adapted to a specific pH range. For some
organisms, the optimum pH range is quite narrow. A sudden or drastic pH change will
kill some of these organisms or reduce survival and reproduction, which disrupts the
food chain and the aquatic community. Lower pH levels also can release heavy
metals from the surrounding bedrock into the water. Once dissolved, these metals can
interfere with the uptake of dissolved oxygen by aquatic organisms.
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Materials
Included in the materials kit for Biological Water Quality Factors
LaMotte Dissolved Oxygen TesTabs™ (in the
small pouches)
LaMotte Coliform TesTabs™ (in the large
glass vials wrapped in bubble wrap)
125 mL collection bottle
Small glass sample vial
Sample comparison chart
Needed from the equipment kit
100 mL graduated cylinder
Thermometer
Test tube rack
Plastic pipettes
Permanent marker
Needed, but not supplied
Aluminum foil
Timer or stopwatch
GPS or GPS-enabled phone (recommended)
Household bleach solution
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2 packs of 4
2
3
Safety
Gloves and safety glasses should
be worn any time you are
dealing with chemicals, including
the TesTabs™. Natural water
samples can contain harmful
contaminates and organisms.
Wear your gloves and safety
glasses during sample collection
as well.
Read all the instructions for this laboratory activity before beginning. Follow the
instructions closely and observe established laboratory safety practices, including
the use of appropriate personal protective equipment (PPE) described in the Safety
and Procedure section.
Safety while performing field work is of greatest importance. Never conduct field work
on your own, especially in remote areas; always take a responsible partner with you.
Prepare and leave a field safety plan, complete with planned survey locations and
expected return time, with a responsible party. Wear appropriate clothing (long pants
and close-toed shoes), sunblock, and insect repellant if necessary. You may be working
in waterways; therefore, rubber boots or waterproof shoes would be optimum if
available. Be familiar with potential hazards in your chosen survey areas. Never enter a
wetland area if there is a flood risk. Please review the Field Work portion of the Lab
Safety Manual or your school’s specific safety guidelines for more information. Make
sure that you are healthy enough to participate in field work. If in doubt, consult your
medical practitioner.
Do not eat, drink, or chew gum while performing this activity. Wash your hands with
soap and water before and after performing the activity. Clean up the work area
with soap and water after completing the investigation. Keep pets and children
away from lab materials and equipment.
Preparation
1. Select your field sites for the following activities. This kit has enough test materials
for two locations. You will perform activities 1, 2 and 4 for your first sample in their
entirety before collecting a second sample at a second site and performing the
activities again. Activities 3 and 5 must be started before collecting your second
sample, but the activites can run concurrently for both samples once started.
2. Ideal water sources include shallow creeks, rivers, springs, or ponds. Locate
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several possible test sites on a map. Make sure these areas are accessible to you.
Sites located in county or state parks are accessible, but you will need to acquire
permission from landowners before visiting privately owned sites.
3. Activities 1 and 2 will be performed at the field site. Activity 3 will be started at
the field site and completed after 5 days of incubation. Activities 4 and 5 can be
started at the field site or within 23 hours of sample collection. It is common
practice to use a cooler to transport samples because they need to be kept
cool and dark to prevent post-sampling photosynthesis from skewing test results.
4. Select an area to test based on the way the watershed flows. Choosing sites
above and below the convergence of two waterways helps pinpoint the source
of any pollution found in the samples. Prepare a map that marks the chosen
sites. Each site can be identified with latitude and longitude using a GPS or GPSenabled phone. For accurate measurement of dissolved oxygen (Biological
Water Quality), determine the approximate altitude of the sample location.
There are several web sites that can help you identify the altitude of a pinned
location (e.g., http://www.freemaptools.com/elevation-finder.htm and
http://www.daftlogic.com/sandbox-google-maps-find-altitude.htm). There are
also free apps for smart phones, such as “My Altitude” for iOS and “Get Altitude”
for Android.
5. Gather all materials that you need before starting the field trip. Make sure that
you have the LaMotte TesTabs™, the water-resistant sample comparison sheets,
Table 1 and Figure 1, the data table, a thermometer, sterile water sample
collection bottles, aluminum foil, gloves, goggles, a marking pen, and a cooler
with ice if you will be transporting samples.
6. At the test location, collect each sample from a location that is representative of
the waterway per the activity instructions. Best results will be obtained if samples
are collected from locations that are not close to storm water drains and
drainage ditches. For the most reliable results, water samples should be
collected midstream. Water near the shore generally is not representative of an
entire stream or river, because some runoff is highly concentrated and has not
yet mixed with the main body of water. In some situations, (e.g., if the water is
too deep), it may not be possible, feasible, or safe to obtain an ideal sample. In
this case, collect a water sample from a safe location.
Activity 1: Temperature Measurement
1. While wearing gloves, submerge the thermometer 4 inches below the surface of
the water.
2. Keep the thermometer in the water for 2 minutes. Remove it from the water and
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immediately record the water temperature at this site in the Data Table.
Activity 2: Sample Collection
1. Remove the lid from the 125 mL sampling container.
2. Rinse the container two or three times with water from the sample site.
3. Replace the lid.
4. Plunge the sample bottle several inches under the surface of the water.
5. Remove the lid.
6. Turn the bottle into the current, if there is one, and allow the water to fill the
container. Hold the sample container underwater for at least 30 seconds.
7. Replace the cap while the bottle is still submerged. There should be no air left in
the bottle.
8. This sample will be used for Activities 45.
Activity 3: Dissolved Oxygen and Biological Oxygen Demand Test
1. This test should be performed at the test site. Wear gloves when working with test
site water.
2. Measure and record the temperature of the water site as described in Activity 1.
3. Submerge a small glass tube several inches under the water surface. Make sure
that the water completely fills the tube. Remove it from the water.
4. Drop two Dissolved Oxygen TesTabs™ into the tube. Water will overflow the tube.
5. Place the cap on the tube and tighten it without allowing any air bubbles into
the tube.
6. Mix the sample by repeatedly turning the tube upside down and righting it until
the tablets are dissolved. This will take approximately 4 minutes.
7. Incubate the sample for 5 more minutes.
8. While sample 1 is incubating, repeat step 3. Cap the tube and tighten it without
allowing any air bubbles into the tube. Wrap the tube in aluminum foil and set
aside at room temperature for 5 days. This is sample 2.
9. After sample 1 has incubated for 5 minutes, compare the sample color with the
color chart. Record the result as ppm dissolved oxygen in the data table
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The remainder of this procedure can be completed at home.
10. Calculate dissolved oxygen percent saturation as follows:
a. To obtain the corrected DO value, multiply the DO (ppm) value by the
correction factor that corresponds to the altitude of the test site. Refer to
the Altitude Correction Factor Table (Table 1).
b. Print out Figure 1. Locate the corrected DO value on the “Oxygen (ppm)”
axis, and mark the point.
c. Mark the temperature of the sample on the “Water temperature (°C)” axis
of Figure 1.
d. Using a straightedge, connect the two points. The intersection of the line
with the “% Saturation” value is the corrected dissolved oxygen percent
saturation level for the sample.
e. Record the DO % Saturation value in the data table.
11. After 5 days, unwrap sample 2 and repeat steps 410.
12. Calculate the BOD by taking the difference between the two samples. BOD is
reported in ppm. Record the BOD in the data table.
©2015,Carolina Biological Supply Company
Table 1: Altitude Correction Factors
Equivalent
Altitude (ft.)
0
542
1,094
1,688
2,274
2,864
3,466
4,082
4,756
5,403
6,065
6,744
7,440
8,204
8,939
9,694
10,472
Figure 1: Dissolved Oxygen Saturation
©2015,Carolina Biological Supply Company
Correction Factor
1
0.98
0.96
0.94
0.92
0.90
0.88
0.86
0.84
0.82
0.80
0.78
0.76
0.74
0.72
0.70
0.68
Activity 4: Turbidity Test
1. Wear gloves when working with test site water.
2. Fill the graduated cylinder from your equipment kit to the 100 mL mark with water
from your collection bottle.
3. Remove the cylinder from its base and place it over the black and white pattern
on the turbidity chart.
4. Look straight down through the water sample, and observe the pattern through
the water column. If you cannot see the pattern, proceed to step 4a. If you can,
proceed to step 4b.
a. Pour off a small amount of water (
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