AP LAB #12 DISSOLVED OXYGEN Introduction to Water Quality & Ms. Gaynor AP Biology

Why is Water SO Important? It is a renewable resource that is becoming polluted & destroyed Recycles through “water (hydrologic) cycle” WATER = SURVIVAL ~ 80% of body = water Only 1% of water on Earth = fresh Used for domestic, industrial, commercial, & recreational use

Why Test Water Quality? To monitor human and ecological impact and its affect on aquatic life/ aquatic habitats Monitoring means “to collect data” on water to access affects of pollutants Water quality can disrupt water chemistry  harmful to aquatic food chain

AP LAB #12: Dissolved Oxygen Dissolved Oxygen (DO) = the amount of oxygen gas dissolved in water 95% more O2 in air than in cold water [DO] used to determine whether the biological activities requiring O2 are occurring ALSO an indicator of pollution O2 important in metabolic processes  indicator of water quality

Why is DO important? Oxygen must be in water in a free state (O2) before organisms can use it This depends on physical and chemical properties of water. Sewage and detritus (dead organic matter) depletes DO b/c it requires a lot of O2 as it decomposes

How does the O2 get into the water? primary productivity (photosynthesis) Atmosphere (air) Winds Mixing (waves, currents etc…) 6

6 MAIN Factors that affect [DO] 1. Air pressure above water affects [DO] Less O2 present at higher elevations (b/c air is less dense) water at higher elevations contain less O2

6 MAIN Factors that affect [DO] (con’t) 2. Photosynthesis (adds O2) & cell aerobic respiration (uses/takes away O2) Photosynthesis in bright light  aquatic producers make more O2 Cellular Respiration (metabolic) activity aquatic organisms use more O2

6 MAIN Factors that affect [DO] (con’t) 3. As salinity and temperature of water increases  [DO] decreases Salinity = amount of salts dissolved in water More Salts= less [DO] Less Salts= more [DO] Temperature Warmer H2O = low [DO] Cold H2O = higher [DO]

6 MAIN Factors that affect [DO] (con’t) 4. Decomposition activity As organic (DEAD) material decays, microorganisms that decompose material consume oxygen (O2) 5. Mixing and turbulence Wave action, waterfalls, and rapids all aerate (add “air” to) water and increase the O2 concentration.

6 MAIN Factors that affect [DO] (con’t) 6. Wind Windless nights = low(er) [DO] High winds = High [DO]

Where is the greatest [DO]? lowest [DO]?

Think: “Ewww” a green lake Think: “Ohhh” a nice clear blue lake DO and Trophic States 2 classifications Eutrophic = “well nourished” high nutrient [ ]  lots of plant life as well as respiration (O2 fluctuates) GREENER in color (from algae) Oligotrophic = low nutrient [ ]  low plant growth (always rich in O2 ) CLEAR in color, very “blue” Think: “Ewww” a green lake Think: “Ohhh” a nice clear blue lake

EUTROPHIC OLIGOTROPHIC

Lab #12: Dissolved Oxygen Concepts dissolved O2 (DO) primary productivity (aka-photosynthesis) measured in 3 ways: amount of CO2 used rate of glucose/ sugar (biomass) formation rate of O2 production net productivity vs. gross productivity cellular respiration 16

Reminder… PHOTOSYNTHESIS Cellular Respiration 6CO2 + 6H2O  C6H12O6 + 6O2 carbon water glucose oxygen gas dioxide (sugar) Cellular Respiration C6H12O6 + 6O2  6CO2 + 6H2O + ATP glucose oxygen gas carbon water energy (sugar) dioxide

Biological Influences & Dissolved Oxygen As photosynthesis increases, oxygen levels increase: CO2 + H2O Biomass + O2 As respiration increases due to decay or organism’s need for ATP increases, oxygen levels decrease: Biomass + O2 CO2 + H2O

Productivity in a Aquatic Environment Primary productivity = rate at which organic (carbon based) materials are stored through photosynthesis Can be measured in three ways: 1. The amt of carbon dioxide used 2. The rate of sugar formation 3. The rate of oxygen production

Productivity gross primary productivity (GPP)= TOTAL Photosynthesis ALL the sugar (biomass) made during photosynthesis Net productivity (NNP) = sugar (biomass) that remains after photosynthesis minus sugar being used up by cellular respiration P– CR = NP net productivity = gross productivity – respiration

Light & Dark Bottle Method Light bottle represent daytime Dark bottle can represent night time BOTTLE #1 (5 light bottles 1 light control bottle) **exposed to light **Acts as initial bottle as well BOTTLE #2 (1 dark bottle) **NOT exposed to light

LIGHT BOTTLES 100%, 65%, 25%, 10%, and 2% 2004-2005 23

What is going on in the Light Bottles (2%, 10%, 25%, 65%, 100%)?

What is going on in the Dark Bottle (foil wrapped)?

You need to isolate variables using the Light & Dark Bottle Method Light Bottle - Dark Bottle = Gross PP +O2 & -O2 -O2 only Total + O2 Net PP = Light Bottle – Initial Bottle +O2 & -O2 starting [DO] Cellular Respiration = Initial – Dark You need to isolate variables using the Light & Dark Bottle Method

Lab #12: Experimental Design (setup) This lab has 2 PARTS and will take 2 days!

Lab #12: Experimental Design (setup) PART A Prepare 3 water sampling bottles filled with pond water Use 3 pond water samples from at different temperatures refrigerator (cold) incubator (warm) room temperature DO NOT FORGET TO TAKE THE TEMPERATURE AT EACH CONDITION; RECORD ON WORKSHEETS! Determine the [DO] of each sample using the Winkler Titration Method. Record your values in your data table Estimate your % saturation using a nomogram Record class results

Nomograph To measure how much O2 the water can hold (saturation), you need to be able to read a nomograph.

Lab #12: Experimental Design (setup) PART B Fill 7 water sampling bottles with pond water (BOD bottles for “biological oxygen demand”) Add piece of plant to each bottle. Make sure that they are the same! Count leaves, measure, etc. CONTROL THE VARIABLES! Label bottles-I, D, 100%, 65%, 25%, 10%, and 2% Determine the DO for the initial bottle using Winkler Titration method. THIS IS YOUR BASE LINE (CONTROL GROUP); the amount of DO ALL your samples start with! Cover D (dark) bottle with aluminum foil Cover the remaining bottles with screens (see procedure for details) Place bottles under the light source. Wait 24 hours, then measure DO levels of remaining samples using Winkler Titration method.

Lab 12: Dissolved Oxygen 2004-2005 32

Hints for Setting Up Lab #12 Rinse sampling bottle 3x with sample water Submerge bottle in water; allow to fill. Tap bottle to release air bubbles You do NOT want extra atmospheric O2 While bottle is submerged, replace cap If there are air bubbles in the bottle, empty and repeat Preserve sample immediately. Test within 2 hours.

More Hints for Setting Up Lab #12 Do NOT leave water samples uncapped! Hello O2 will get in!!! Use rubber bands to secure the screens NOTE: We are using HORNWORT plant not Anacharis Please FIX on your worksheets!

Winkler Titration Method (PART 1) How Do You FIX Samples Correctly? To “fix” your samples, follow for each sample bottle: (“fix” = trap DO in sample) Uncap bottle Add 8 drops of MgSO4 (manganous sulfate) to bottle (can cause cancer/stain) Add 8 drops of alkaline potassium iodide azide to bottle (can cause cancer/stain) Cap bottles and mix. A precipitate will form! Allow precipitate to settle to shoulder of bottle. Add a spoonful (1 g) of sulfamic acid powder to bottle (STRONG ACID) Cap and mix. Precipitate should dissolve!

Winkler Titration Method (PART 2) How Do You Determine the amount of DO in the samples Correctly? To determine the [DO] of your samples, follow for each sample bottle, including the initial bottle: Uncap bottle Carefully fill the titration tube/cup to the 20 mL line. Fill the titration syringe to the “0” (zero) line with sodium thiosulfate. Add 1 drop AT A TIME to sample and swirl after each drop. Do this until you get a FAINT yellow color. Remove titration syringe Add 8 drops of starch indicator solution Swirl the sample. The sample should now be BLUE! No blue = no measurable [DO] or too much sodium thiosulfate 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. Read titration syringe scale for result in mg DO/liter Fill in data table; multiple DO in mg/L by 0.698 to convert to mL/L

Chemical Reactions To Preserve DO: Done in the field FIX To Preserve DO: Done in the field O2 + 2 Mn2+ + 2H2O 2Mn(IV)O2 + 4H+ (pH >10) Allow precipitate to settle (reaction goes to completion) 2Mn(IV)O2 + 4H+ + 2I- Mn2+ + I2 (yellow) + 2H2O (low pH) DO is preserved (“fixed”) -------------------------------------------------------------------------- To Test Sample: This step can be done in the lab. Na2S2O3 + 4I2 + 5H2O 8I- + 2SO42- + 10H+ + Na+ (the titration) Starch + I2 blue (to improve endpoint determination) SAMPLE

Amt of Thiosulfate Used = Amt of DO in water Titrate w/ Thiosulfate 20 mL STARTING POINT MIDDLE POINT Very DARK yellow PALE yellow Add 8 drops of STARCH Turns BLUE PART 1 PART 2

ADD ALL THIOSULFATE USED FROM STARTING POINT TO END POINT! CONTINUE to Titrate w/ SAME 20 mL Thiosulfate END POINT Turns CLEAR!!! ADD ALL THIOSULFATE USED FROM STARTING POINT TO END POINT! This is your amount of thiosulfate used = amount of DO in water Each # = 1 mL = 1 ppm = 1 mg/L 1 mg/L x 0.698 = 1 mL/L 1 2 3 4 5 6 7 8 9 10

Dissolved Oxygen Can be measured in absolutes (how much O2 is actually in the water) or by % saturation % saturation - measure of DO compared to how much DO COULD be in the water Water at a higher temperature CAN NOT hold as much oxygen as cold water (use nonmaogram)

Lab 12: Dissolved Oxygen Conclusions to Think About…  temperature =  dissolved O2  light =  photosynthesis =  O2 production O2 is consumed during cellular respiration  respiration =  dissolved O2 (consumption of O2) 42