AP Biology



AP Biology - AP Lab #13: PRE-LAB INFORMATION Ms. Day

AP Lab #13: Dissolved Oxygen & Primary Productivity

Introduction

In an aquatic environment, oxygen must be in a solution in a free state (O2) before it is available for use by organisms (bio-available). Its concentration and distribution in the aquatic environment are directly dependent on chemical and physical factors and are greatly affected by biological processes. In the atmosphere, there is an abundance of oxygen, with about 200 mL of oxygen/1 L air. In an aquatic environment, there are about 5-10 mL O2/1 L water. The concentration of the oxygen in aquatic environments is a very important component of water quality.

At 20(C, oxygen diffuses 300,000 times faster in air than water, making the distribution of oxygen in air relatively uniform. Spatial distribution of oxygen in water, on the other hand, can be highly variable, especially in the absence of mixing by currents, winds, and tides.

Other chemical and physical factors – such as salinity, pH, and especially temperature- can affect the dissolved oxygen (DO) concentration and distribution. Salinity, usually expressed as parts per thousand (ppt), is the content of dissolved salts in water. Generally, as temperature and salinity increase, the solubility of oxygen in water decreases.

The partial pressure of oxygen in the air above the water affects the amount of DO in the water. Less oxygen is present at higher elevations since the air itself is less dense; therefore the water at high elevations contains less oxygen. At 4,000 meters in elevation (13,000 feet), the amount of dissolved oxygen in water is less than two-thirds what it is at sea level. All of these physical factors, along with oxygen concentration, work together to increase diversity in aquatic habitats.

Oxygen from the atmosphere is mixed into the water through diffusion. However, more oxygen is mixed into the water with the help of winds, rain, waves, and currents. The faster the water moves, the more dissolved oxygen the water will contain since it has more contact time with the air. The process of photosynthesis (underwater plants and algae) occurring in the water affects the number and kinds of animals found there. Healthy streams are saturated with oxygen (90 to 110% saturation) during most of the year.

Biological processes, such as photosynthesis and respiration, can also significantly affect DO concentration. Photosynthesis usually increases the DO concentration in water. Aerobic respiration requires oxygen and will usually decrease DO concentration. The measurement of the DO concentration of a body of water is often used to determine whether the biological activities requiring oxygen are occurring and consequently, it is an important indicator of pollution.

The primary productivity of an ecosystem is defined as the rate at which organic materials (carbon-containing compounds) are stored. Only those organisms possessing photosynthetic pigments can utilize sunlight to create organic compounds from simple inorganic substances. Green plants obtain carbon for carbohydrate synthesis from the carbon dioxide in the water or the air according to the basic equation for photosynthesis:

6CO2 + 6 H2O ( C6H12O6 + 6O2

The rate of carbon dioxide utilization, the rate of formation of organic compounds, or the rate of oxygen production can be used as a basis for measuring primary productivity. A measure of oxygen production over time provides a means of calculating the amount of carbon that has been bound (taken up by photosynthesis) over a period of time. For each mL of oxygen produced, approximately 0.536 mg of carbon has been assimilated.

One method of measuring the rate of oxygen production is the light and dark bottle method. In this method, the DO concentrations of samples of ocean, lake, or river water or samples of laboratory algal cultures are measured and compared before and after incubation in light and darkness. The difference between the measurements of DO in the initial and dark bottles is an indication of the amount of oxygen that is being consumed in respiration by organisms in the bottle. In the bottles exposed to light, the biological process of photosynthesis and respiration are occurring; therefore the change over time in DO concentration from the initial concentrations is the measure of net productivity. The difference over time between the DO concentrations in the light bottle and the dark bottle is the total oxygen production and therefore an estimate of gross productivity.

AP Lab #12: Dissolved Oxygen & Primary Productivity

Procedure

NOTE: Sample data is using Chlorella (scientific name= Chlorella pyrenoidosa), which is unicellular green alga. You are using Anarcharis (aka-Elodea; scientific name= Egeria Densa)

Part A: Dissolved Oxygen and Temperature:

1 ppm (Each tiny tick mark (line) between the numbers = 0.2 ppm, which equals 1 drop)

5 drops= 1 ppm = 1 mg/L

mg/L ( mL/L= multiply by 0.698

F.Y.I( Stoichiometry

1 molecule of oxygen = 4 molecules of ppt ( ppt molecules dissolve after acid into yellow/brown color

Therefore, [DO] is directly proportionate to the yellow/brown color. Darker color = more DO in sample. No blue after starch indicator= no measurable DO. It can also mean too much thiosulfate titrate before adding 8 drops of starch indicator.

Part A (Day 1): Dissolved Oxygen and Temperature

1. Fill three of the water sampling bottles with water of the three different temperatures provided.

2. Using the Winkler Method Handout, determine the amount of DO per sample.

3. Your own nomograph for oxygen saturation is attached. Use a straight edge or ruler to determine the percent DO in each of your temperature samples. (To use a nomograph, you line up the edge of a ruler with the temperature of the water on the top scale and the DO on the bottom scale, and read the percent saturation on the middle scale.)

4. Record the temperature, DO, class mean DO, your group % Saturation, and Class mean % saturation in the data table below. To figure out the % saturation, you need to use the nomogram (see below) and a ruler.

Table 13.1: Temperature/DO Data

|Temperature (ºC) |Lab Group DO (ppm) |Class Mean DO (ppm) |Lab Group % saturation |Class Mean % Saturation |

| | | | | |

| | | | | |

| | | | | |

Nomogram (note: use this nomogram NOT the one in the College Board lab manual, which is incorrect)

[pic]

How to use this ‘thing’:

For a quick and easy determination of the percent saturation value for dissolved oxygen at a given temperature, use the saturation chart above. Pair up the mg/l of dissolved oxygen you measured and the temperature of the water in degrees C using a ruler. Again, using a ruler, draw a straight line between the water temperature and the mg/l of dissolved oxygen. The percent saturation is the value where the line intercepts the saturation scale. Record the % saturation for all three temperatures in your data table above (ALSO IN YOUR NOTEBOOK!!!)

Note: Streams with a saturation value of 90% or above are considered healthy.

Percent Saturation values of 80 - 120% are considered to be excellent, and values less than 60% or over 125% are considered to be poor.

REMEMBER to collect QUALITATIVE data:

• Do NOT forget to record qualitative data, including descriptive. sketches with labels, etc. for PART A. These need to be included in your formal lab report.

Part B (Day 1): A Model of Productivity as a Function of Depth in a Lake

1. Obtain 7 water-sampling bottles. Using tape, label the cap of each bottle. Label: I (for initial), D (for dark), 100%, 65%, 25%, 10%, 2%.

2. Fill all of the bottles COMPLETELY with the lake water provided.

3. Determine the DO for the “Initial” bottle ONLY right now using the Winkler Titration Method (see below) and record its value in the data table below (see Table 12.2 below). This is the amount of DO that the water has to start with (baseline). This baseline vial (initial bottle) is your control group. All vials should have the SAME baseline DO value to start out with.

▪ We will fill in the rest of Table 12.2 on Day two. We will also calculate a class mean.

Table 13.2: Respiration

| |Individual Data (ppm) |Class Mean (ppm) |

|Initial DO | | |

|Dark Bottle DO | | |

|Respiration Rate (Initial-Dark) | | |

Winkler Titration Method

To “fix” your samples, follow for each sample bottle:

1. Uncap bottle

2. Add 8 drops of MgSO4 (manganous sulfate) to bottle (can cause cancer/stain)

3. Add 8 drops of alkaline potassium iodide azide to bottle (can cause cancer/stain)

4. Cap bottles and mix. A precipitate will form! Allow precipitate to settle to shoulder of bottle.

5. Add a spoonful (1 g) of sulfamic acid powder to bottle (STRONG ACID-BE CAREFUL!)

6. Cap and mix. Precipitate should dissolve!

7. Your sample is now “fix” meaning no more O2 that enters or leaves will have an effect on your DO reading/titration.

To determine the [DO] of your samples, follow for each sample bottle, including the initial bottle:

8. Uncap bottle

9. Carefully fill the titration glass vial to the 20 mL line. Be as accurate as possible!

10. Fill the titration syringe to the “0” (zero) line with sodium thiosulfate.

11. Add 1 drop AT A TIME to sample and swirl after each drop. Do this until you get a FAINT yellow color. Use a piece of WHITE paper under your titrating vial to gauge when you are at the FAINT yellow color.

12. Remove titration syringe

13. Add 8 drops of starch indicator solution

14. Swirl the sample. The sample should now be BLUE!

▪ No blue = no measurable [DO] or too much sodium thiosulfate!

15. Add sodium thiosulfate 1 drop at a time and SWIRL until BLUE color disappears. If you finish the syringe and it is still blue, fill syringe again and continue. Kept track of how many syringe full’s you use!

16. Once the blue color is gone, you are finished titrating. Read the titration syringe scale for your DO concentration data in mg DO/liter.

17. Fill in data table with data ; multiple DO in mg/L by 0.698 to convert to mL/L.

4. Cover the “Dark” bottle with aluminum foil so that no light can enter.

▪ In this bottle, photosynthesis can NOT occur, so the only things that will change DO will be the process of cellular respiration by all of the organisms present.

5. The decrease of natural light that occurs due to depth in a body of water will be simulated by using window screen. Wrap screen layers around the bottles in the following way:

• 100% light – no screens; 65% light – 1 screen layer; 25% light –3 screen layers; 10% light –5 screen layers; and 2% light- 8 screen layers.

• The bottles will be on their sides under the lights, so remember to cover the bottom of the bottles to prevent light from entering there.

• Use rubber bands to keep the screens in place.

6. Place the bottles on their sides under the bank of lights in the classroom. Tomorrow we will continue…

Part B (Day 2): A Model of Productivity as a Function of Depth in a Lake

1. To complete this experiment, determine the DO in all of the bottles that have been under the lights as well as the DO in the dark bottle.

2. Record the Dark Bottle Do in Table 12.2 (from Day One).

3. Calculate the Respiration rate (Initial ppm-Dark ppm) and record in Table 12.2 (from Day One).

4. Use the “dark” DO in the Table 12.2 to calculate gross productivity in Table 12.3 (see below).

5. Record the DO values for other bottles.

6. Complete the calculations to determine the Gross and Net Productivity for each bottle. The calculations will be based on a time period of one day (24 hours).

7. Determine class means and record in Table 12.4 (see below).

8. Multiply all DO values in mg/L (which is the same as ppm) by 0.698 to convert to mL/L. Record values for your individual group data in Table 12. 5 and values for class mean data in Table 12. 6 (see tables below on pg. 4).

Table 13.3 Individual Group Data: Productivity of Screen-Wrapped Sample

|# of screens |% light |DO |Gross Productivity (ppm) |Net Productivity (ppm) |

| | |(ppm) |[light bottle – dark bottle] |[light bottle – initial bottle] |

|0 |100 | | | |

|1 |65 | | | |

|3 |25 | | | |

|5 |10 | | | |

|8 |2 | | | |

Table 13.4 Class Mean Data: Productivity of Screen-Wrapped Sample

|# of screens |% light |DO |Gross Productivity (ppm) |Net Productivity (ppm) |

| | |(ppm) |[light bottle – dark bottle] |[light bottle – initial bottle] |

|0 |100 | | | |

|1 |65 | | | |

|3 |25 | | | |

|5 |10 | | | |

|8 |2 | | | |

REMEMBER to collect QUALITATIVE data:

• Do NOT forget to record qualitative data, including descriptive. Sketches with labels, etc. for PART B. These need to be included in your formal lab report.

GRAPHS FOR PART B (in Results section of your formal lab report):

• In your formal lab writeup, you will need to graph both net and gross productivities as a function of light intensity for your class mean data ONLY.

o This will be Figure 2 in your formal lab report.

• In your formal lab writeup, you will also need to graph the concentration of DO (dissolved oxygen) in mL/L as a function of light intensity for your class mean data ONLY.

o This will be Figure 3 in your formal lab report.

• Make sure these LINE graphs are properly labeled and neatly done.

• ALWAYS MAKE LINE GRAPHS with a key, title and labeled axes. Excel works well when making printable graphs for your formal lab reports.

Pre-lab

AP Lab #13: Dissolved Oxygen and Aquatic Primary Productivity

1. Instructions (do at home):

• Read the information above (introduction and procedure) and prepare for a pre-lab quiz.

• Please read/study the virtual lab instructions and animations for this lab at

o At the end of the virtual lab on this website for Lab #13, take the “Self Quiz”

o After you complete it, click “check answers”… If you got any wrong, please study your mistakes.

• READ THE PROCEDURE CAREFULLY!!! Focus on the follow:

• Problem/Question for Lab #13:

▪ What is the primary productivity of a lake or stream? What is the effect of changing light intensity (lake depth) on primary productivity?

• Hypotheses:

▪ PART A: You need 1 hypothesis

▪ PART B: You need 2 different hypotheses here; 1 for DARK bottle and 1 for LIGHT bottles (collectively). These hypotheses should be in the “If…then…because…” format, please.

• Independent Variable: What are you testing or changing in the experimental groups?

• Dependent Variable: What are you measuring? HINT: what are the numerical data representing?

• Control Group: Know the bottle being used as the control group and why

• Procedure including procedure notes, hints, etc. (if applicable)

• Data Tables: Know the data tables that are being used for this lab

AP Biology - AP Lab #13: PRE-LAB INFORMATION Ms. Day

Name: _________________________________________ Date: ____________ Period: ______

AP Biology- Lab #12: Dissolved Oxygen (Student Rough Draft Worksheets)

Part A (Day 1): Dissolved Oxygen and Temperature

Qualitative Data:

Quantitative Data:

Table 12.1: Temperature/DO Data

|Temperature (ºC) |Lab Group DO (ppm) |Class Mean DO (ppm) |Lab Group % saturation |Class Mean % Saturation |

| | | | | |

| | | | | |

| | | | | |

Nomogram (note: please use this nanogram to determine the % saturation for your lab group/class mean)

[pic]

Part B (Day 1): A Model of Productivity as a Function of Depth in a Lake

Qualitative Data:

Quantitative Data:

Table 12.2: Respiration

| |Individual Data (ppm) |Class Mean (ppm) |

|Initial DO | | |

|Dark Bottle DO | | |

|Respiration Rate (Initial-Dark) | | |

Table 12.3 Lab Group Data: Productivity of Screen-Wrapped Sample

|# of screens |% light |DO |Gross Productivity (ppm) |Net Productivity (ppm) |

| | |(ppm) |[light bottle – dark bottle] |[light bottle – initial bottle] |

|0 |100 | | | |

|1 |65 | | | |

|3 |25 | | | |

|5 |10 | | | |

|8 |2 | | | |

Table 12.4 Class Mean Data: Productivity of Screen-Wrapped Sample

|# of screens |% light |DO |Gross Productivity (ppm) |Net Productivity (ppm) |

| | |(ppm) |[light bottle – dark bottle] |[light bottle – initial bottle] |

|0 |100 | | | |

|1 |65 | | | |

|3 |25 | | | |

|5 |10 | | | |

|8 |2 | | | |

Winkler Titration Method

To “fix” your samples, follow for each sample bottle:

1. Uncap bottle

2. Add 8 drops of MgSO4 (manganous sulfate) to bottle (be careful…can cause cancer/stain)

3. Add 8 drops of alkaline potassium iodide azide to bottle (be careful…can cause cancer/stain)

4. Cap bottles and mix. A precipitate will form! Allow precipitate to settle to shoulder of bottle.

5. Add a spoonful (1 g) of sulfamic acid powder to bottle (STRONG ACID-BE CAREFUL!)

6. Cap and mix. Precipitate should dissolve! Your sample is now “fix” ( O2 that enters or leaves will have an effect on your DO reading.

To determine the [DO] of your samples: follow the directions below for each sample bottle, including the initial bottle:

7. Uncap bottle

8. Carefully fill the titration glass vial to the 20 mL line. Be as accurate as possible!

9. Fill the titration syringe (green and pink in color) to the “0” (zero) line with sodium thiosulfate.

10. Add 1 drop AT A TIME to sample and swirl after each drop. Do this until you get a FAINT yellow color. Use a piece of WHITE paper under your titrating vial to gauge when you are at the FAINT yellow color.

11. Remove titration syringe and set aside. DO NOT EMPTY THIS SYRINGE!!!

12. Add 8 drops of starch indicator solution

13. Swirl the sample. The sample should now be BLUE! (No blue = no measurable [DO] or too much sodium thiosulfate!)

14. Add sodium thiosulfate (FROM THE SAME SYRINGE IN STEP #11) 1 drop at a time and SWIRL until BLUE color disappears. If you finish the syringe and it is still blue, fill syringe again and continue. Kept track of how many syringe full’s you use (i.e.-how much sodium thiosulfate is used ALL TOGETHER FROM STEP #1 until now)!

15. Once the blue color is gone, you are finished titrating. Read the titration syringe scale for your DO concentration data in mg DO/liter.

16. Fill in data table with data ; Remember 5 drops = 1 ppm of DO = 1 mg of DO/1 L of H2O. Each syringe holds 10 ppm’s.

-----------------------

L(ight)

Net Productivity

DO2 (ml O2/L)

I(nitial)

Gross

Productivity

Respiration

D(ark)

L – I = Net Productivity

I – D = Respiration

L - D = Gross Productivity (or, think of it as L – I + I – D)

0 Incubation time (hours) 24

1 2 3 4 5 6 7 8 9 10

LIGHT BOTTLES

DARK BOTTLE (FOIL WRAPPED)

**NO PHOTOSYNTHESIS

[DO] = is the amount of O2 used by the plant in CELLULAR RESPIRATION

Cellular Respiration =

INITIAL BOTTLE – DARK BOTTLE

Gross Primary Productivity (GPP) =

LIGHT BOTTLE (100%, 65%, 25%, 10% or 2%) - DARK BOTTLE

(Photosynthesis + Cellular Respiration) – Cellular Respiration

Net Primary Productivity (NP) =

LIGHT BOTTLE (100%, 65%, 25%, 10% or 2%) – INITIAL BOTTLE

(Photosynthesis + Cellular Respiration) – Original Amount of DO

Photosynthesis – cellular respiration

INITIAL BOTTLE

(CONTROL)

**NO CELLULAR RESPIRATION

**This is the STARTING AMOUNT OF DO in ALL 7 bottles!

2% LIGHT BOTTLE

**NO/VERY LITTLE PHOTOSYNTHESIS

+

CELLULAR RESPIRATION OCCURING

65% LIGHT BOTTLE

**SOME PHOTOSYNTHESIS

+

CELLULAR RESPIRATION OCCURING

10% LIGHT BOTTLE

**VERY LITTLE PHOTOSYNTHESIS

+

CELLULAR RESPIRATION OCCURING

25% LIGHT BOTTLE

**LITTLE PHOTOSYNTHESIS

+

CELLULAR RESPIRATION OCCURING

100% LIGHT BOTTLE

**LOTS OF PHOTOSYNTHESIS

+

CELLULAR RESPIRATION OCCURING

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