Materials Included In Kit

Additional Materials Required

Prelab Preparation

1. Prepare a 0.002% solution of bromthymol blue by mixing 20 mL of the supplied 0.04% bromthymol blue solution and 380 mL of distilled water.
2. Make sure this solution is blue. If it is not, add dilute sodium hydroxide (0.1 M) dropwise until it is blue.
3. Prepare enough (1 L) 80 °C and 45 °C DI water for five student groups. Also have 500 mL of room temperature DI water on hand for student use.

Safety Precautions

Wear chemical splash goggles, chemical-resistant gloves and a chemical-resistant apron. Wear heat-resistant gloves when handling the beakers with the heated water. Wash hands thoroughly with soap and water before leaving the laboratory. Follow all laboratory safety guidelines. Please review current Safety Data Sheets for additional safety, handling and disposal information.

Disposal

Please consult your current Flinn Scientific Catalog/Reference Manual for general guidelines and specific procedures, and review all federal, state and local regulations that may apply, before proceeding.The resulting solutions may be disposed of according to Flinn Suggested Disposal Method #26b.

Teacher Tips

• Enough materials are provided in this kit for 5 groups of students. This laboratory activity can reasonably be completed in one 50-minute class period.
• As an extention, calcium sulfate may be used to show that it will not inhibit gas production or prevent a color change in the indicator. Iron(III) sulfate will inhibit the CO₂ gas production and the indicator in the sample tube should remain blue.
• One hot plate station may be used if desired so that the heated distilled water solutions required for every student group may be dispensed at the same time.
• This is a great activity to perform during discussions about respiration, fermentation and environmental conditions.
• If enough hot plates are not available, an option for hot water is to use a large hot pot or coffee perculator. A laboratory microwave is another option. Be sure to use beaker tongs or wear heat-resistant gloves.
• If sucrose is used instead of dextrose, it will take longer to begin producing CO₂, but the rate will eventually be the same as the flasks containing dextrose. Yeast cells are able to begin respiration more quickly using a monosaccharide (e.g., dextrose) than a disaccharide (e.g., sucrose) as a food source. This is due to the inability of yeast to metabolize disaccharides. In order for yeast to use sucrose as an energy source, the yeast must first break it down into its component disaccharides of glucose and fructose—these are called invert sugars. This extra step, which delays the start of fermentation, is carried out by the enzyme invertase.
• Ammonium oxalate, calcium chloride and iron(III) sulfate will all affect the production of gas and a color change in the indicator, with iron(III) sulfate preventing these occurrences altogether. Iron, halide ions and cyanide ions also have the greatest ability to inhibit or prevent yeast metabolism. Even NaCl in high enough concentration will prevent metabolism in yeasts.
• The following is a list of common ions (and their sources) found in water sources:

  Carbonate: Comes from the high water solubility of carbonate material and rocks, such as limestone and dolomite. Another source of carbonate ions is from dissolved atmospheric carbon dioxide. When heated in the presence of calcium and magnesium, alkaline water containing carbonates will form crust-like scales in pipes restricting the flow of fluids and also causing the release of carbon dioxide gas.

  Chloride: Dissolved from rocks and soils and present in sewage and industrial wastes. When chloride is in excess of 100 parts per million, water has a salty taste and when it is in excess of 150 parts per million, physiological damage may occur. Chloride also increases the corrosiveness of water when it is present in higher concentrations.

  Chromate: Comes from industrial waste and is used to prevent corrosion in cooling towers. When chromate accumulates in large amounts in the human body it may cause cancer.

  Iron: Present in natural sources, such as rocks and soils. Iron is also found in industrial waste and is present from the corrosion of iron pipes. When the concentration of iron is above 100 parts per billion, it will precipitate when it comes in contact with air. This will in turn stain laundry, plumbing fixtures, and silverware. It will also cause food and drinks to taste and look peculiar.

  Lead: Found in water from plumbing, coal, and gasoline sources. A major source of lead pollution came from lead additives in gasoline prior to the 1980s (the structure of the gas has since been changed to cut down on the pollution risk). The EPA estimates that 20% of our exposure to lead comes from drinking water. At blood levels greater than 100 parts per billion, anemia, kidney damage and mental retardation can result.

  Phosphate: Comes from fertilizers, detergents, wastewater of domestic origin such as human, animal and plant residue, and from wastewaters of industrial origin. Phosphate is added to farm and city water systems to control hardness. Phosphates from laundry detergents can result in overgrowth of algae, which in turn will cause the algae to die at a high rate and undergo decomposition. This decomposition process depletes oxygen from the water and will result in increased fish kill.

  Sulfate: Dissolved from rocks and soils that contain sulfides and gypsum. Sulfate is also due to industrial waste in liquid or gaseous forms. When sulfate is present with calcium, scale will result in pipes and steam boilers. Sulfate concentrations of 500 parts per million produce water that tastes bitter and sulfate concentrations of 1,000 parts per million may be cathartic.

Sample Data

Temperature and Yeast

Answers to Questions

1. How did the temperature of the solutions affect the yeast cultures? The yeast culture at 45 °C produced the most CO₂ and was the most viable.

   The yeast culture at room temperature eventually produced CO₂ but at a much slower rate than the 45 °C solution. The yeast culture at 80 °C did not produce any CO₂.

2. Given the results, describe the relationship among the 80 °C, 45 °C and room temperature solutions. Include the relationship between dissolved oxygen and CO₂ production of the yeast.

   Initially, as the temperature increased the rate at which the yeast consumed oxygen increased. The increased temperature decreased the ability of the oxygen to dissolve in water. This means that the yeast used up the available oxygen more quickly at the higher temperatures. If the temperature was too hot (80 °C), the yeast cells were not viable.

3. How did the iron(III) sulfate effect the yeast growth and CO₂ production?

   The iron(III) sulfate eliminated the CO₂ production and viability of the yeast culture.

4. What happened when the containers were swirled at each five-minute mark? Why?

   For the containers that produced CO₂, the amount of gas travelling from the container to the sample tube increased. More oxygen was introduced to the containers as they were swirled.

5. A dam has just been installed in a local river. How would the dissolved oxygen readings above and below the dam vary?

   The water below the dam should be oxygen rich due to the increased motion of the water. The water above the dam should be more stagnant and therefore would be more likely to have less oxygen than the water below.

6. A nuclear power plant with a cooling lake is being installed in your area. The water from the cooling lake is eventually going to be introduced to a local natural water source. What effect would the water from the cooling lake have on aquatic organisms if the water from the power plant was introduced to the natural water source before it completely cooled?

   Answers will vary. It is really dependent on the temperature and amount of the cooling lake water entering the natural source and the overall size of the water source.

7. Describe two ways pollution could be reduced or prevented in natural water sources in your area.

   Student answers will vary.

Introduction

Illustrate the effects of temperature and chemicals on the ability of yeast to metabolize sugar. This activity can be used to simulate the effects of chemical pollution/improper disposal on any type of organism.

Concepts

• Chemical pollution
• Thermal pollution
• Dissolved oxygen

Background

Water is an essential natural resource. It is used for almost every activity in life and is required for life itself. Water is needed for agricultural and industrial use, drinking, transportation and in recreation. Water often seems to be available in an almost endless supply, but as populations rise and our world becomes increasingly industrialized, more and more water is being utilized. With this extensive use of water, a problem arises: The water becomes polluted and contaminated. This pollution leads to a strain on water’s ability to recycle and cleanse itself of contaminants. The amount of water available and its distribution and quality are critical issues that affect all life. An increasing awareness of the need to monitor the quality of water and to locate the sources of water pollution is essential to protecting this vital resource.

Thermal pollution is the degradation of water quality by any process that changes ambient water temperature. A common cause of thermal pollution is the use of water as a coolant by power plants and industrial manufacturers and from storm water discharged to surface waters from roads and parking lots. When water used as a coolant is returned to the natural environment at a higher temperature, the change in temperature impacts organisms by decreasing oxygen supply and affecting ecosystem composition.

Dissolved oxygen (DO) is the amount of gaseous oxygen, O₂, dissolved in a body of water. Oxygen enters into the water by aeration, diffusion from air and as a byproduct of photosynthesis. In general, high flow rates or water turbulence will increase oxygen levels in water due to aeration. Slow moving or stagnant water usually has very low oxygen levels. Oxygen levels also change throughout the day as a result of photosynthesis, usually peaking in late afternoon. The amount of oxygen that will dissolve in water depends on temperature and pressure and is very sensitive to environmental conditions.

Dissolved oxygen is inversely related to temperature—as the water temperature increases, the amount of oxygen that can dissolve decreases. In the summer, extremely warm water temperatures may result in very low dissolved oxygen. Typical environmental factors that affect DO levels include the amount of organic matter or waste from decaying vegetation, the presence of nitrates, phosphates and other nutrients and the concentration of electrolytes, such as Na⁺, Ca²⁺, Mg²⁺, Cl^– and HCO₃^– ions.

Dissolved oxygen is one of the most important indicators of the overall health of a body of water. When water contains a large amount of oxygen (8–10 ppm), the quality of the water is generally very good. Water with consistently low dissolved oxygen levels (<3–4 ppm) is extremely stressful to aquatic organisms and may harbor only a few species adapted to such conditions. DO levels less than 2 ppm will not support fish life.

Thermal pollution may also increase the metabolic rate of aquatic organisms. an increased metabolic rate may result in food source shortages, causing a sharp decrease in a population. A temperature change of even one to two degrees may cause significant changes in organism metabolism and other adverse cellular biology effects. principal adverse changes may include reduction of permeability of cell walls and coagulation of cell proteins. These cellular level effects can negatively affect reproduction and increase mortality rates.

In this activity, yeast will be used as a model organisms to demonstrate the effect of both temperature and chemical pollution on living organisms. Yeast cells are facultative anaerobes, that is, they can carry out either aerobic or anaerobic respiration. In the absence of oxygen, yeast will carry out alcoholic fermentation—they will ferment simple sugars to ethyl alcohol and CO₂. Although yeasts are considered to be hardy organisms, the presence of particular conditions will prevent their respiration.

Experiment Overview

In this activity, a pH indicator solution known as bromthymol blue will be used to study the viability of yeast cultures under different environmental conditions. During an “incubation” period, the yeast cultures will be observed to see if gas bubbles form and if the indicator changes color. Carbon dioxide dissolves in water to give an acidic solution (carbonic acid). Bromthymol blue solution is blue when the pH is >7.6 and yellow when the pH is <6.

Materials

Safety Precautions

Wear chemical splash goggles, chemical-resistant gloves and a chemical-resistant apron. Wear heat-resistant gloves when handling the beakers with the heated water. Wash hands thoroughly with soap and water before leaving the laboratory. Follow all laboratory safety guidelines.

Procedure

Part I. Temperature and Yeast

1. Obtain four containers with holes in the lid, four pieces of 12" tubing, four water sample tubes, yeast, dextrose and bromthymol blue solution.
2. Place the four sample tubes into a test tube rack.
3. Place 10 mL of the bromthymol blue solution into one of the sample tubes.
4. Repeat step 3 for the three other sample tubes.
5. Place one end of the 12" plastic tubing into the hole of one of the container lids. Be sure that the tip of the tube just barely extends into the end of the cap (see Figure 1).
   {12016_Procedure_Figure_1}
6. Repeat step 5 for the three other container lids.
7. Using a balance and weighing dish, weigh 2 g of yeast.
8. Place the 2 g of yeast into one of the 120-mL containers.
9. Repeat steps 7 and 8 using the other three 120-mL containers.
10. Using a balance and weighing dish, weigh out 1 g of dextrose.
11. Place the 1 g of dextrose into one of the 120-mL containers.
12. Repeat steps 10 and 11 for the other three 120-mL containers.
13. Using a balance and a clean weighing dish, weigh out 3 g of iron(III) sulfate. Place the 3 g of iron(III) sulfate into one of the containers that already contains the 2 g of yeast and 1 g of dextrose.
14. Obtain 100 mL of 80 °C DI water, 200 mL of 40 °C DI water and 100 mL of room temperature DI water from the instructor.
15. Place another 100 mL of distilled water into the first plastic container. Measure and record the temperature on the worksheet.
16. Pour 100 mL of 45 °C DI water in the second and fourth [the container with iron(III) sulfate] containers and 100 mL of room temperature DI water in the third container as shown in Figure 2.
    {12016_Procedure_Figure_2}
17. Immediately screw on the caps of the containers.
18. Swirl each container and submerge the free end of the tube into the bromthymol blue solution in each sample tube (see Figure 2).
19. Record the current time on the worksheet.
20. Record all observations every 5 minutes for a total of 30 minutes. Swirl each container every 5 minutes as well. If the bromthymol blue solution begins to travel up the plastic tubing and towards the container with the yeast, simply tap the tubing so the bromthymol blue solution returns to the sample tube.
21. Answer the Post-Lab Questions for Part I.
22. Consult your instructor for appropriate disposal procedures.

Student Worksheet PDF

12016_Student1.pdf