Fermentation of Glucose by Yeast: Lab Explained

Introduction

Glucose → Ethanol + Carbon dioxide

In the absence of oxygen, enzymes from microorganisms break down sugars through the chemical process called fermentation. Because they possess distinctive sets of metabolic genes, microorganisms like bacteria and fungi can develop enzymes that can break down various kinds of sugar compounds. Therefore, the flavor of fermented food can change significantly when the types and numbers of bacteria are altered. Certain cells in our body without mitochondria use the mechanism of fermentation.

Specifically, in adulthood, our red blood cells lose their mitochondria. The citric acid cycle and redox reactions along the electron transport chain occur in mitochondria. Our blood’s mature red blood cells use lactic acid fermentation to produce chemical energy; this makes sure that none of the oxygen that the red blood cells transport is used by them. Moreover, Fermentation can take one of three fundamental kinds. There are three types of fermentation: Lactic acid, Ethanol, and Acetic acid. An overview of testing for the presence of ethanol is provided in this experiment.

During the Lactic acid fermentation process, Lactic acid and other waste products are produced through this anaerobic process by the breakdown of starches or sugars. In addition to preventing food from microbial deterioration, lactic acid bacteria also shield the body from microbial disease. Fermentation using lactic acid is frequently used to make foods like yogurt, pickles, and sauerkraut. Alcoholic fermentation is one of the oldest and most significant fermentation processes used in the biotechnology industry.

Alcoholic fermentation occurs in the yeast’s cytoplasm without oxygen (Sablayrolles, 2009; Stanbury et al., 2013). However, it is typically carried out with the yeast Saccharomyces cerevisiae due to its hardiness with low pH and high ethanol tolerance. The regeneration of the NAD+ used during glycolysis is done; this gives yeast a boost of two ATP molecules through the metabolism of hexose. In this type of fermentation, yeasts convert the pyruvate molecules in carbohydrates into alcohol and carbon dioxide molecules, along with bread and beer.

Although the bacterium Zymomonas mobilis has a higher specific ethanol productivity and yield from glucose and sucrose, there are essentially two elements to this process; In the process of glycolysis, glucose is split into two pyruvate molecules. Pyruvate molecules undergo fermentation, changing into two molecules of carbon dioxide and two molecules of ethanol. In addition to ethanol, ethanol fermentation also produces additional substances such esters, higher alcohols, succinic acid, glycerol, 2,3-butanediol, diacetyl, and acetoin. Acetic acid fermentation is an oxidation process that turns carbohydrates from fruits or grains into vinegar and condiments with a sour flavor.

Under aerobic circumstances, acetic acid bacteria (AAB) may oxidize ethanol as a substrate to create acetic acid in neutral and acidic media. AAB are highly tolerant to hop bitterness compounds and can survive in high ethanol concentrations (Priest, 2006). AAB can develop uninvitedly and impart an unfavorable sour flavor to other fermented beverages like wine, cider, beer, functional drinks, and soft drinks because it oxidizes ethanol to acetic acid. Other metabolic products these bacteria produce include gluconic acid, l-sorbose, and bacterial cellulose, which have potential uses in the food and biomedical industries. Based on morphological, physiological, and genetic traits, the categorization of AAB into various genera has undergone a number of changes in recent years.

Materials

• Glass jar with cap
• Table sugar
• Yeast
• Warm water
• Balloons

Method

• All materials were gathered and set up.
• Warm water, followed by table sugar, was added to the bottle and then swirled to dissolve the sugar.
• After the sugar had been dissolved in the warm water, a packet of yeast was slowly added to the bottle.
• In the bottle containing the sugar, water, and yeast mixture, additional swirling was done to aid in the dissolution of the yeast.
• The bottle’s entire aperture was then carefully covered by stretching a balloon over it and onto its neck.
• The process was left to sit in a war for several minutes, and observations were recorded.

Results

Observations

During the fermentation process, the formation of bubbles were seen, and the yeast in the mixture began to rise as the balloon on the bottle began to increase in size, as time passed, a larger inflation was seen, and the balloon eventually fell. When the balloon was removed from the bottle, it had the appearance of a foamy layer at the top of the bottle with numerous bubbles, a liquid portion at the bottom, and the combination smelled strongly of alcohol and was very sour.

Questions

• Is fermentation affected by temperature? Explain.

Yes.

This is so because the temperature has an impact on the speed of chemical reactions, and it directly affects yeast fermentation. The yeast will produce the most carbon dioxide when it is at its ideal temperature, which is 67°C. However, yeast will create more carbon dioxide as the temperature rises, but eventually, when the yeast cells have become denatured due to the temperature rise, the output of carbon dioxide will start to decline.

• What substance, other than ethanol, is produced during fermentation?

Carbon Dioxide.

• Describe an appropriate control that you could have used in this experiment.

An appropriate control that could have been used in the experiment would be the type of sugar used, because the type of sugar used impacts how quickly yeast produces alcohol.

• How did you know when the fermentation was finished?

The researcher knew the process was complete after noticing that the gas emissions halted because the balloon had stopped expanding.

• Name a carbohydrate that you supplied to the yeast as an energy source.

The carbohydrate supplied to the yeast as an energy source was table sugar.

• Give an account of a chemical test to demonstrate that alcohol has been produced.

A chemical test that can be used to demonstrate that alcohol has been produced is the Jones test which uses chromium trioxide in the presence of sulfuric acid to act as a potent oxidizing agent. A primary alcohol initially becomes an aldehyde when Jones’ reagent is present. The secondary alcohol will next undergo oxidation, turning it into a ketone, while the carboxylic acid will follow. The key to this test is the chromium oxidation state. In Jones’ reagent, chromium is in the +6 oxidation state. The reagent’s vibrant crimson and orange color is a result of the presence of Cr(VI) complexes.

• Why does fermentation eventually cease? 

Fermentation eventually ceases because there is insufficient sugar left to finish the reaction because the yeast cells consume all of the sugar or because of a rise in the alcohol concentration, destroying the yeast cells.

Discussion

A fungus called yeast breaks down sugar into CO₂ and ethanol as a byproduct. Carbon dioxide gas is created as the sugar is consumed by the yeast. Inside the balloon, the trapped CO₂ gradually builds up and causes it to inflate. The balloon inflated after a few minutes because a reaction occurs when particles meet with energies greater than or equal to the activation energy. Active yeast interacted with the sugar to make ethanol and CO₂ gas, inflating the balloon. However, because there was so much sugar added and because the rate of fermentation increased as sugar concentration increased, particle collisions increased, and the fermentation process produced considerably more byproducts, which encouraged the active yeast to produce more CO2 and ethanol. As a result, the balloon expanded slightly before inflation stopped.

Conclusion

In conclusion, yeast will ferment in the presence of sugar to produce ethanol, carbon dioxide, and energy. As such, the type of sugar used in the process impacts how quickly yeast produces alcohol.

References

1. Fully Integrated Lab Management Systems are Key to Removing Lab Bottlenecks that Hinder Quality. (n.d.). Lab Manager. https://www.labmanager.com/in-focus/fully-integrated-lab-management-systems-are-key-to-removing-lab-bottlenecks-that-hinder-quality-28267
2. Biology Online. (2019, October 7). Fermentation Definition and Examples – Biology Online Dictionary. Biology Articles, Tutorials & Dictionary Online. https://www.biologyonline.com/dictionary/fermentation
3. Alcoholic Fermentation – Definition, Location, Equation, Products, Reactants, Agent, Subproducts. (n.d.). BYJUS. https://byjus.com/neet/alcoholic-fermentation/
4. Ethanol Fermentation – an overview | ScienceDirect Topics. (2010). Sciencedirect.com. https://www.sciencedirect.com/topics/agricultural-and-biological-sciences/ethanol-fermentation
5. Acetic Acid Bacteria – an overview | ScienceDirect Topics. (2011). Sciencedirect.com. https://www.sciencedirect.com/topics/agricultural-and-biological-sciences/acetic-acid-bacteria
6. Gomes, R. J., Borges, M. de F., Rosa, M. de F., Castro-Gómez, R. J. H., & Spinosa, W. A. (2018). Acetic Acid Bacteria in the Food Industry: Systematics, Characteristics and Applications. Food Technology and Biotechnology, 56(2). https://doi.org/10.17113/ftb.56.02.18.5593