BIOL240 Lecture Notes - Lecture 18: Bromocresol Purple, Hydrogen Sulfide, Gut Flora

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30 Oct 2017

School

University of Waterloo

Department

Biology

Course

BIOL240

Professor

Cheryl Duxbury

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Document Summary

Experiment 18: some metabolic activities of bacteria: fermentation of carbohydrates: Can produce acidic, alcoholic, or gaseous end products. Can be distinguished by the carbohydrates they do or do not utilize, as well as the products formed. Often prepared as tube broths, each contain a small inverted fermentation tube (or. Durham tube) for trapping gas formed when broth is inoculated and cultured. Each broth contains basic nutrients, one specific carbohydrate, and colour indicator (bromocresol purple is used) to indicate ph if acid is produced. Organisms that grow in broth but do not use carbohydrate does not change the colour of the medium and no gas will be found. Organisms may produce acid products but no gas. Some organisms that do not ferment may produce alkaline end products by utilizing protein nutrients and will also change colour: production of indole: By-product of metabolic breakdown of amino acid tryptophan. If indole is present it combines with reagent to produce brilliant red.

Related Questions

Experiment 1 Fermentation by Yeast Experiment Inventory Labware (4) 250 mL Beakers (1) 100 mL Graduated Cylinder (1) Test Tube Rack (5) Fermentation Tubes = (10) Test Tubes (5 plastic and 5 glass; see Figure 4) (1) Measuring Spoon (4) Pipettes (1) Ruler Note: You must provide the materials listed in *red. EXPERIMENT 1: FERMENTATION BY YEAST Yeast cells produce ethanol, C2 H6 O, and carbon dioxide, CO2 , during alcoholic fermentation. In this experiment, you will measure the production of CO2 to determine the rate of fermentation in the presence of different carbohydrates with fermentation tubes. Note: Regular table sugar is sucrose, a disaccharide, which is made up of glucose and fructose. Glucose is a monosaccharide. Figure 4: Fermentation tubes. Note how the smaller, plastic test tube is inverted into the larger glass tube. You will create five fermentation tubes in this experiment. PROCEDURE 1. In this experiment, you will mix yeast with sugar, EqualÂ®, and SplendaÂ®. Before you begin, develop a hypothesis predicting what will happen when the sugar/sweeteners are mixed with yeast. Will fermentation occur? Why or why not? Record your hypothesis in the post-lab questions. 2. Use the permanent marker to label three 250 mL beakers as EqualÂ®, SplendaÂ®, and Sugar. 3. Empty the EqualÂ®, SplendaÂ®, and Sugar packets into the corresponding beakers. 4. Fill the EqualÂ® and SplendaÂ® beakers to the 100 mL mark with warm tap water. 5. Fill the Sugar beaker to the 200 mL mark with warm tap water. 6. Mix each beaker thoroughly by pipetting the solution up and down several times. Use a new pipette to mix each solution. Each beaker now contains a 1% solution. Set these aside for later use. 7. Completely fill one of the smaller plastic tubes with tap water and invert the larger glass tube over it. Push the small tube up into the larger tube until the top connects with the bottom of the inverted tube. Invert the fermentation tube (Figure 4) so that the larger tube is upright (there should be a small bubble at the top of the internal tube). Note: Repeat Step 7 several times as practice. Strive for the smallest bubble possible. When you feel comfortable with this technique, empty the test tube(s) and proceed to Step 8. CAUTION: Do not try to force the plastic test tube into the glass test tube. This might cause your glass test tube to break, causing you injury. If your plastic test tubes do not fit easily, please call eScience Labs for replacement glass tubes. If you are able to set up at least two fermentation tubes, continue with the experiment, but know that you will have to perform steps 12-15 in multiple steps. 8. Use the permanent marker to label the fourth 250 mL beaker as Yeast. 9. Fill this beaker with 175 mL of warm tap water. It should be between 30 and 40o C (warm to the touch). 10.Open the yeast package, and use the measuring spoon to measure and pour 1 tsp. yeast into the beaker. Pipette the solution up and down until all of the yeast is mixed homogenously into the solution. Note: Make sure the yeast solution remains homogenous before each test tube is filled in the proceeding steps. The yeast density is fairly high, and the yeast may settle to the bottom of the beaker if it rests for an extended period of time. 11. Use the permanent marker to label the big glass and small plastic test tubes as 1, 2, 3, 4, and 5. 12.Use the 100 mL graduated cylinder to measure and pour 15 mL of the following solutions into the corresponding small plastic test tubes: Tube 1: 1% Glucose Solution Tube 2: 1% Sucrose Solution Tube 3: 1% EqualÂ® Solution Tube 4: 1% SplendaÂ® Solution Tube 5: 1% Sugar Solution Note: Thoroughly rinse the graduated cylinder between each measurement. 13.Fill the remaining volume in each small tube to the top with the yeast solution. 14.Slide the corresponding larger tube over the small tube and invert it as practiced in Step 7. This will mix the yeast and sugar/sweetener solutions. 15.Place the fermentation tubes in the test tube rack, and use a ruler to measure (in millimeters) the initial air space in the rounded bottom of the internal tube. Record these values in the Table 1. 16.Allow the test tubes to sit in a warm place (approximately 30 Â°C) for two hours. Placement suggestions include: a sunny window sill, atop (not in!) a warm oven heated to approximately 85 Â°C (185 Â°F on an oven setting), or under a very bright (warm) light. 17.At the end of the fermentation period, use your ruler to measure (in millimeters) the final gas height (total air space) in each tube. Record this data in Table 1. 18.Calculate the difference between the initial and final gas height in each tube. Record this data in Table 1.

EXPERIMENT 1: FERMENTATION BY YEAST

Result Tables

Table 1: Yeast Fermentation Data

Tube	Initial Gas Height (mm)	Final Gas Height (mm)	Net Change (mm)
1
2
3
4
5

Post-Lab Questions

Include your hypothesis from Step 1 here. Be sure to include at least one piece of scientific reasoning in your hypothesis to support your predictions.

Did you notice a difference in the rate of respiration between the various sugars? Did the artificial sugar provide a good starting material for fermentation?

Was anaerobic fermentation occurring? How do you know (use scientific reasoning)?

If you observed respiration, identify the gas that was produced. Suggest two methods you could use for positively identifying this gas.

Hypothesize why some of the sugar or sweetener solutions were not metabolized, while others were. Research the chemical formula of Equal^Â® and Splenda^Â® and explain how it would affect yeast respiration.

How do the results of this experiment relate to the role yeast plays in baking?

What would you expect to see if the yeast cell metabolism slowed down? How could this be done?

Indicate sources of error and suggest improvement (for example, what types of controls could be added?).

BIOL240 Lecture Notes - Lecture 18: Bromocresol Purple, Hydrogen Sulfide, Gut Flora

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