Lab 10: Glycolysis and Fermentation

What’s so sweet about Sugar?

Objectives• Discuss ATP and where the ‘energy’ is

• Describe what fermentation is and where the ‘energy’ is

• Design an experiment to ‘test’ fermentation

• Recognize/explain what happens in a RedOx reaction

Thinking it Through• CH4 (methane)

C2H5OH H3C-CH2-OH (ethanol) C8H18 CH3(CH2)6CH3

(octane)

• H2O CO2

• Similarities in components/structure?

• Differences in behavior?

• Differences in structure?

So where are ‘calories’?

Energy (heat) needed to raise 1g water, 1oC

Saturated Fat

RedOx Reactions

OIL RIG

OIL = Oxidation Is Loss

RIG = Reduction Is Gainof electrons

of electrons

ATP

ATP

ATP

ATP

Metabolic Pathways

Metabolic Pathways

Metabolic Pathways

Glycolysis• Glyco – glucose/sugar

• Lysis – “to loosen”, take apart

Glycolysis

Freeman, Scott. Biological Science. 4th Edition.

Glycolysis + fermentation

Freeman, Scott. Biological Science. 4th Edition.

Glycolysis + Fermentation

Freeman, Scott. Biological Science. 4th Edition.

Here’s the claim:Glycolysis –

glucose + 2ADP + 2Pi + 2NAD+ => 2 pyruvic acid + 2NADH+ + 2ATP

Here’s the claim:Glycolysis –

Fermentation –

glucose + 2ADP + 2Pi + 2NAD+ => 2 pyruvic acid + 2NADH+ + 2ATP

pyruvic acid + NADH+ => ethanol + CO2 + 2NAD+

Here’s the claim:Glycolysis –

Fermentation –

What can you test?

Rank in terms of feasibility, safety, etc.

glucose + 2ADP + 2Pi + 2NAD+ => 2 pyruvic acid + 2NADH+ + 2ATP

pyruvic acid + NADH+ => ethanol + CO2 + 2NAD+

Here’s the claim:Glycolysis –

Fermentation –

What can you test?

Rank in terms of feasibility, safety, etc.

glucose + 2ADP + 2Pi + 2NAD+ => 2 pyruvic acid + 2NADH+ + 2ATP

pyruvic acid + NADH+ => ethanol + CO2 + 2NAD+

Here’s the claim:Glycolysis –

Fermentation –

What can you test?

Rank in terms of feasibility, safety, etc.

What will glyco + ferment?

How????

glucose + 2ADP + 2Pi + 2NAD+ => 2 pyruvic acid + 2NADH+ + 2ATP

pyruvic acid + NADH+ => ethanol + CO2 + 2NAD+

Standard Protocol

• Pg. 10-7 of lab manual

• Change to 500 mL

Here’s the claim:Glycolysis –

Fermentation –

What can you test?

Rank in terms of feasibility, safety, etc.

What will glyco + ferment?

How/when to do CO2, sugar, ETOH?

glucose + 2ADP + 2Pi + 2NAD+ => 2 pyruvic acid + 2NADH+ + 2ATP

pyruvic acid + NADH+ => ethanol + CO2 + 2NAD+

Sugar/CO2 Presence?

• Appendix C

• ***READ APPENDIX C*** – REALLY (lots of helpful stuff…in the text)

• What about ETOH?

Group discussion

• On what basis will you be drawing conclusions at the end of the experiment– making comparisons between the final result

and...?

• What if we wanted to produce a lot of ETOH, or very little glucose?

Optimizations• Concept: limiting components. How do you

max your yield (sensitivity) for:– Glucose (how might you limit in final solution?)

– CO2 (Maximize)

– ETOH (Maximize)

• Use standard protocol (10-7) as a baseline and adjust amounts from there per your role (could there be too much bread?)

Optimizations• Concept: limiting components. How do you

max your yield (sensitivity) for:– Glucose (how might you limit in final solution?)

– CO2 (Maximize)

– ETOH (Maximize)

• Use standard protocol (10-7) as a baseline and adjust amounts from there per your role (could there be too much bread?)

• What do we do? Glucose groups – 1X or 2X?

After Mixing

• What will your solution look like?– “murky”?

• How do you speed it up?

• What conditions do we WANT?

First part

• 1) Yeast mixed with glucose solution• 2) Take glucose pre-sample (in plastic tube)

and place on ice – how much? Answer: see ‘candidate solution’ in protocol

• 3) Hook up BaOH glass tube to flask and follow protocol

• 4) Cook after sealing/taping• Turn on hot plates

After Cooking

• 1) BaOH outcome?• 2) Take another yeast solution extract for

sugar testing – how many do we have? Why?– How to get chunks out?– Follow Benedict’s protocol

• 3) Heat the beer…get ETOH – how to do so?– How to test?

no heating parafilm!!!

Thermometer should measure GAS temp, leave it up high!

AFTER

What did we prove?

• Consistent with…

Next Week• Assess: Glycolysis and fermentation

• Lab report: Fermentation with a Bullet– One/group in Dropbox