Fermentation 1

Alcoholic + Industrial Applications

Define respiration (aerobic).

An energy-yielding process that occurs in the presence of oxygen as final electron acceptor.

What is an example of aerobic respiration?

The complete catabolism of sugar (glucose) to CO2 + H2O and much energy.

What is anaerobic respiration?

Occurs in the absence of oxygen. Not all energy is extracted from glucose and an intermediate metabolite, such as lactic acid, is produced.

What happens to lactic acid if oxygen is re-supplied?

Lactic acid breakdown.

In microorganisms, what is anaerobic respiration referred to as?

Fermentation

Define fermentation.

A catabolism that doesn’t utilise oxygen, but the organic compounds involved are both the electron donor and acceptor. Energy generation often involves substrate-level phosphorylation.

Common electron donors in fermentation?

Sugars

Common electron acceptors in fermentation?

• Pyruvate

• Acetaldehyde

• Fumarate

• Protons

• Acetyl-CoA

What is the role of pyruvate?

Yield lactate and/or opines.

What is the product when acetaldehyde acts as an electron acceptor?

Ethanol.

What is the product when fumarate acts as an electron acceptor?

Succinate.

What is the product when protons (H⁺) act as electron acceptors?

Hydrogen gas (H₂).

What is the product when acetyl-CoA acts as an electron acceptor?

Fatty acids and derivatives.

What organism produces beer and spirits?

Yeast

What organisms produce wine and spirits?

Yeast and fungi

What organisms produce bread?

Yeast and bacteria.

What products does lactic acid bacteria produce (4)?

• Fermented meats

• Sauerkraut, kimchi

• Yoghurt, sour cream

• Olives

What organism produces vinegar?

Acetic acid bacteria.

How does fermentation help preserve food?

Change pH,

deplete nutrients,

produce inhibitors (e.g., acid, ethanol)

How does fermentation improve sensory attributes?

Produce, release, modify compounds to alter flavour/aroma, taste, etc.

What are the health benefits of fermentation?

Breakdown complex molecules to aid digestion

degrade toxins,

produce beneficial compounds

support probiotic microbes.

How does fermentation improve sustainability?

Valourize food waste into useful by-products.

What valuable by-products can fermentation produce?

Enzymes,

biomass,

pigments,

biofuels,

chemicals,

vaccines, etc.

What is the basic equation for alcoholic (ethanolic) fermentation?

Fermentation is an (1) process that acts on (2) sugars (monosaccharides).

1. Intracellular

2. Simple

It begins with uptake of (3). (4) sugars require breakdown first.

3. Sugars

4. Complex

Uptake is ordered due to (5).

Energetic efficiency.

How are Glu and Fru uptaken?

Facilitated diffusion.

How is Mal uptaken?

Active uptake + maltase (enzyme)

How is Suc broken down? What enzyme is used?

Broken down to glucose + fructose by invertase.

What happens to the sugars after uptake?

Feed into glycolysis.

What does glycolysis ultimately end with?

Pyruvate.

Draw process of glycolysis.

GLYCOLYSIS

What does 1× 6-carbon Glu convert to?

2× 3- carbon pyruvate.

What does 2× 3-C G3P convert to?

2× 3-C pyruvate.

How much ATP and NADH is produced in this process?

4 ATP

2 NADH

In presence of O2 and through TCA cycle and ETC, what is pyruvate metabolised to?

CO2 and H2O.

In presence of O2, and through TCA cycle and ETC, how muchATP is produced by NADH/FADH2 when re-oxidised?

36 ATP.

In absence of O2, what is used to re-oxidise NADH?

Fermentation

During fermentation, what is pyruvate converted to?

Acetaldehyde

What is re-oxidised furing fermentation?

Ethanol and NADH

How much ATP is yielded via fermentation?

2

What is the Pasteur effect?

Cell growth increases with aeration.

What is the Crabtree effect?

Tendency of some yeast (Saccharomyces cerevisae) to ferment despite the presence of oxygen.

What is the main difference between white and red wine?

The inclusion of skins (provide colour and flavour in reds).

Draw the wine fermentation process.

Why is completion of fermentation to produce wine challenging?

• High osmolarity (sugars)

• High ethanol

• Low oxygen

• Low nutrition

BIOETHANOL

How much petroleum is used globally per day?

104.3 million barrels/day.

What is the percentage of ethanol to gasoline in E5, E10 and E85 gasoline replacement?

E5= 5%

E10= 10%

E85=85%

What are the 3 largest producers on bioethanol?

US, Brazil, China

Name 3 biowastes.

Straw

Food waste

Surplus wine

Name 5 sugar-containing feedstock considerations.

Sugar beet

Sugar cane

Molasses

Whey

Sweet Sorghum.

Name 3 starch-containing feedstock considerations.

Corn

Wheat

Root crops (cassava)

Name 4 lignocellulosic biomass feedstock considerations.

Straw

Agricultural waste

Crop

Wood residues

How are sugar-rich plants processed?

Milling or dilution of molasses.

Sucrose→ Glu + Fru

How is whey (4.9% lactose) processed?

B-galctosidase to produce galactose and glucose.

How is starch processed?

Mix of linear (amylose) and branched (amylopectin) polyglucans.

a-amylase, active on a-1,4 (not a-1,6) linkages in amylopectin.

Use a-amylase and glucoamylase → glucose syrup.

What is lignocellulose?

Cross-linked carbohydrate polymers of cellulose (30-50%) and hemicellulose (20-40%) with long non-carbohydrate lignin (10-20%).

What is needed before fermentation of lignocellulose?

Pretreatment and enzymatic hydrolysis.

What are the challenges of lignocellulose fermentation?

Pretreatment and energy cost

Toxicity of breakdown products

Non-utilisation of xylose

What are the physical methods for lignocellulose pretreatment?

Milling, freezing, microwave, extrusion.

What are the chemical methods for lignocellulose pretreatment?

Acid, alkaline, oxidative, ionic liquid, organosolv, ozonolysis (to promote enzymatic digestibility).

What are the physico-chemical methods for lignocellulose pretreatment?

Steam explosion, liquid hot water, wet oxidation, CO2 explosion, ammonia fibre explosion, and ultrasonication.

What inhibitors are formed during lignocellulose pretreatment?

Hi/Lo pH,

MF,

formic acid,

acetic acid,

levulinic acid,

phenolic compounds

How can inhibitors from lignocellulose pretreatment be removed?

Membrane methods,

activated carbon, or

using resistant yeast.

What are the main sugars in lignocellulose composition?

Glucose (60–70%) and xylose (30–40%)

How does Saccharomyces cerevisiae perform with xylose?

poorly

Which yeast can use xylose, and how efficiently?

Yeast like Scheffersomyces (Pichia) stipitis can use xylose but not vigorously.

What are potential solutions to improve xylose utilization?

Genetic modification (GM) of yeast strains.

What is required for glucose and xylose fermentation in genetically modified Saccharomyces?

Introduction of genes from S. stipitis: XR (xylose reductase) and XDH (xylitol dehydrogenase).

What are the by-products of xylose fermentation when using XR and XDH?

Excess NADH leads to glycerol and xylitol as by-products.

What is an alternative to XR and XDH for xylose fermentation in GM Saccharomyces?

XI (xylose isomerase).

What are the advantages and disadvantages of using XI (xylose isomerase)?

Advantages: Higher ethanol production. Disadvantages: Slower fermentation process.