Ch 13 Carbohydrate Fermentation - Micro Lab

Front 1

carbohydrates

Back 1

the most common energy source for microorganisms

Front 2

chemoheterotrophic organisms

Back 2

obtain energy from 2 basic processes:

1. fermentation
2. respiration

Both catabolic systems convert the chemical energy of organic molecules to high-energy bonds in ATP

Front 3

What are the 3 pathways that can produce energy during respiration?

Back 3

1. glycolysis
2. Krebs cycle
3. oxidative phosphorylation

Front 4

How does respiration differ from fermentation?

Back 4

• requires an external electron acceptor for substrate oxidation

Front 5

aerobic respiration

Back 5

when molecular oxygen is the terminal electron acceptor

Front 6

anaerobic respiration

Back 6

when an inorganic molecule serves as the external terminal electron acceptor and becomes reduced

ex: nitrate (NO₃⁻) or sulfate (SO₄²⁻)

Front 7

fermentation

Back 7

• a biooxidative process that does not require oxygen in which electrons from glucose are accepted by one or more organic substrates
• begins w/ the production of pyruvate via glycolysis (Embden-Meyerhof pathway)
• end product of pyruvate fermentation depends on the specific organism (alcohol, organic acids; lactic, formic, acetic)
• some organisms may release gases such as hydrogen or CO₂

Front 8

carbohydrate fermentation

Back 8

includes the hydrolysis of disaccharides prior to the fermentation reaction

fermentation begins with the production of the intermediate compound pyruvate

Front 9

lactose fermenter

Back 9

an organism that is able to synthesize the enzyme β-galactosidase which splits lactose into the monosaccharides glucose + galactose

Front 10

Biooxidative pathways using glucose as a substrate

Back 10

Glucose is catabolized during glycoloysis (E-M pathway) > 2 Pyruvic acid + 2 NADH + 2 ATP

The pyruvic acid is used in one of 3 pathways:

• Aerobic Respiration > Krebs Cycle > Electron Transport Chain > largest qty of ATP (36)
• Anaerobic Respiration > inorganic ions serve as final electron acceptor
• Fermentation: organic substrate serves as the final electron acceptor > acid or alcohol (+/- gas)

Front 11

fermentation tube

Back 11

used to determine if a bacterium is able to ferment a particular CHO

it contains:

• Durham tube
• Phenol Broth Red (PRB)
• CHO that is being tested (Glucose, Sucrose, Lactose)

Front 12

Durham tube

Back 12

inverted tube inserted into PRB media of the fermentation tube

used to test for gas production during fermentation; you DO NOT know what type of gas was produced

Front 13

Phenol Broth Red (PRB)

Back 13

fermentation media containing a specific CHO (glucose, sucrose, or lactose)

base media contains:

• peptone
• pH indicator - Phenol Red

Front 14

Phenol Red

Back 14

pH indicator used in PRB media

yellow = acidic pH below 6.8

red = in between

pink = alkaline pH above 7.4

Front 15

What does a color change of medium yellow in PRB media indicate?

Back 15

• positive for acid production
• acid production from fermentation of the specific CHO lowers the pH
• this may be accompanied by gas evolution in the Durham tube

Front 16

What does a color change of medium pink in PRB media indicate?

Back 16

• bacteria was NOT able to ferment the specific CHO instead used peptone as an energy source
• peptone > amino acids undergo oxidative deamination to ketoamino acids > ammonia (NH₃)

Front 17

Describe the results of the PRB result in Tube A

Back 17

medium yellow = positive for acid production

some turbidity

gas was produced

fermentation of CHO occurred

Front 18

Describe the results of the PRB result in Tube B

Back 18

medium yellow = positive for acid production

some turbidity

NO gas was produced

fermentation of CHO occurred

Front 19

Describe the results of the PRB result in Tube C

Back 19

uninoculated control

Front 20

Describe the results of the PRB result in Tube D

Back 20

pink = acid production may have occured

alkaline byproducts > CHO was consumed; peptones were catabolized to amino acids > NH₃

some turbidity

NO gas was produced

fermentation of CHO occurred

* this result can occur if the culture is incubated for more than 48 hrs and the CHO are used up

Front 21

Describe the results of the PRB result in Tube E

Back 21

medium pink = alkaline reaction

peptones were catabolized to amino acids > NH₃

some turbidity

NO gas was produced

NO fermentation of CHO occurred

Front 22

What will happen if a PRB tube that undergoes CHO fermentation is incubated for more than 48 hours?

Back 22

The result may be a false negative for acid production. I may be pink as a result of alkaline (basic) byproducts, from peptone catabolism, that were produced AFTER ALL of the CHO was consumed

Front 23

What species of bacteria were used in this experiment?

Back 23

• Alcaligenes faecalis
• Escherichia coli
• Staphylococcus aureus
• Proteus vulgaris