Day 10 part 2 biorefinery, anaerobic digestion, biopolymer

Environmental Biotechnology

Anaerobic Digestion — Fun & Complete Theoretical Summary

(Only the theoretical parts, nothing operational/experimental omitted)

1. What Anaerobic Digestion Is

Anaerobic digestion (AD) is a microbial-mediated process where microbes break down organic matter in the complete absence of oxygen.

Main product:

Biogas = methane (CH₄) + carbon dioxide (CO₂)
Trace gases: H₂S, H₂, etc.

Why methane matters:

It contains the energy value of the biogas.
CO₂ can also be used today for chemical production.

Key trait:

AD is highly versatile: it works at many scales, with many substrates, and in many reactor styles—from high-tech European reactors to rural dome systems in India or black-bag systems in the Andes.

2. What You Can Feed Into an Anaerobic Digester

AD can digest almost anything containing organic matter. Examples include:

Wastewater sludge
Biowaste
Food industry waste
Pharmaceutical waste
Animal manure
Agricultural residues
Slaughterhouse waste
Algae

BUT: Some wastes need pretreatment, and you can improve digestion by mixing substrates (“co-digestion”). Example:

Manure = high nitrogen → risk of ammonia inhibition
Crop residues = high carbon, low nitrogen → Mixing them gives microbes a balanced diet.

3. Possible End Products of AD

AD is not only about methane.

Products include:

Biogas → heat & electricity, or upgraded to biomethane for grid/fuel use
Digestate → nutrient-rich fertilizer
Volatile fatty acids (VFAs) → if the process is intentionally stopped early (this is “AD without biogas”) for chemical production

AD is often placed at the center of biorefinery systems because it accepts diverse organic residues and can integrate with other value-chains.

4. Assessing Whether a Substrate Is Good: Biochemical Methane Potential (BMP) Tests

BMP tests measure:

Biodegradability
Maximum methane yield
Degradation rate (fast vs slow substrates)

How a BMP test works:

Add inoculum (microbes)
Add substrate
Seal to create anaerobic conditions
Mix and measure methane production over time Results give curves showing total methane yield and how long it takes.

Why rate matters: A substrate might have high potential but be too slow to degrade → not profitable.

Methods:

Volumetric systems with CO₂ traps
Automated systems
Manometric systems + gas chromatography

Common sources of error:

Inactive inoculum
Poor substrate storage (already fermented)
Methodological biases (standardized via Shiny app)

5. Substrate Chemistry: TS & VS

Units often reported as:

Nm³ CH₄/kg VS (Nm³ = gas volume at 0°C, 1 atm)

Key definitions:

TS (Total Solids): Dry matter (after drying at 105°C)
VS (Volatile Solids): Organic fraction (burned off at 550°C)

VS represents the biodegradable organic content, so methane yields are compared per kg VS, not per kg of raw waste. Example: Sewage sludge is mostly water → unfair to compare per kg wet weight.

General rule:

Carbohydrates → lowest energy content
Fats → highest energy content

6. Microbiology of Anaerobic Digestion: Four-Step Food Web

AD is a division-of-labor system. No single microbe performs the whole process. The four classical stages:

Hydrolysis
Acidogenesis (fermentation)
Acetogenesis
Methanogenesis

6.1 Hydrolysis 🧬

Breaks down complex polymers → monomers. Handled by many bacteria producing extracellular enzymes.

Often rate-limiting because:

Substrate inaccessible (e.g., lignocellulose)
Insufficient enzymes

Solution: Pretreatments

Mechanical (grinding)
Physical (ultrasound)
Thermal
Chemical (acids/alkalis)
Enzymatic/biological

6.2 Acidogenesis / Fermentation ⚡

Fastest step. Converts monomers → VFAs, ethanol, other products. Microbes gain most of their energy here.

Challenges:

If faster than later steps → VFA accumulation → reactor souring
Hard to predict product spectrum (pH, salinity, conditions change pathways)

If the process is intentionally stopped here, you get anaerobic fermentation platforms for VFA production.

6.3 Acetogenesis 🔁

Converts VFAs (propionate, butyrate) → acetate + H₂. These bacteria are sensitive to conditions and rely on methanogens.

Key concept: Syntrophy Acetogens produce H₂ → increases partial pressure → inhibits themselves. Hydrogenotrophic methanogens must consume the H₂ to keep the reaction thermodynamically possible. They often grow physically close.

Direct interspecies electron transfer (DIET) is possible when conductive particles are added.

If acetoclastic methanogens are absent: Acetate accumulates → special organisms perform syntrophic acetate oxidation (SAO) to produce H₂ + CO₂, which methanogens can use.

6.4 Methanogenesis 🌿🔥

The final, crucial step. Exclusively performed by Archaea, not bacteria.

Two main pathways:

1. Acetoclastic Methanogenesis

Acetate → CH₄ + CO₂

Dominant in many digesters (e.g., sewage sludge)
Performed by Methanothrix and some Methanosarcina
Sensitive organisms

2. Hydrogenotrophic Methanogenesis

H₂ + CO₂ → CH₄

More robust
Many genera can perform this

If acetoclastic methanogens die, the system can shift toward the hydrogenotrophic route using SAO. Too much ammonia, VFAs, or sulfide can inhibit methanogens → process failure.

7. Environmental and Operational Context

Although you asked for theoretical content only, this part is still theoretical (not engineering procedures).

Temperature regimes:

Psychrophilic (4–20°C): slow
Mesophilic (20–40°C): standard; 35–37°C sweet spot
Thermophilic (50–60°C): faster conversion, pathogen reduction, but less stable

Hydraulic Retention Time (HRT) & Solid Retention Time (SRT)

Define how long microbes have to complete digestion.

Organic Loading Rate (OLR)

How much substrate per day. Too high → overload; too low → poor gas yield.

These parameters help choose the right reactor type and operating strategy.

Complete Summary in One Sentence

Anaerobic digestion is a versatile, multi-step microbial ecosystem where complex organic matter is hydrolyzed, fermented, acetogenically converted, and finally methanogenically reduced to methane, with performance governed by substrate chemistry, syntrophic interactions, and thermodynamic constraints.

Quiz

Score: 0/30 (0%)

Q0. Which statement best defines anaerobic digestion?

A microbial process that requires oxygen to break down organic matter into CO₂.

A microbial-mediated process that decomposes organic matter without oxygen and produces biogas.

A purely chemical reaction where heat converts organic matter into methane.

A photosynthetic pathway for converting CO₂ into methane.

Q1. What are the two major components of biogas produced during anaerobic digestion?

Methane and nitrogen

Carbon monoxide and methane

Methane and carbon dioxide

Hydrogen and oxygen

Q2. Why is co-digestion used in anaerobic digestion operations?

To decrease microbial diversity and stabilize pH.

To balance nutrient ratios by mixing substrates with complementary carbon and nitrogen contents.

To eliminate the need for inoculum.

To ensure only a single substrate type is digested at once.

Q3. What does the Biochemical Methane Potential (BMP) test measure?

The rate of oxygen consumption during digestion.

The maximum methane yield and biodegradability of a substrate.

The total microbial biomass in the digester.

The amount of CO₂ retained by the inoculum.

Q4. Why must gas volumes in BMP tests be reported under 'normal' conditions?

Because biogas expands unpredictably at low pressure.

To standardize gas measurements across temperatures and pressures.

To correct for methane solubility in water.

To comply with EU fertilizer regulations.

Q5. What does Total Solids (TS) represent in substrate characterization?

The mineral content after combustion.

The organic matter burned off at 550°C.

The dry matter remaining after heating at 105°C.

The total nitrogen present in the sample.

Q6. Why is hydrolysis often the rate-limiting step in anaerobic digestion?

Methanogens outcompete hydrolytic bacteria.

Complex polymers like lignocellulose are difficult for enzymes to access.

Acetate levels inhibit hydrolysis.

Hydrolysis requires oxygen, which is scarce.

Q7. Which statement about acidogenesis is correct?

It is typically the slowest step in anaerobic digestion.

It produces methane directly from acetate.

It converts monomers into volatile fatty acids and other fermentation products.

It requires hydrogenotrophic methanogens to proceed.

Q8. What is the major challenge acetogens face during acetogenesis?

Competition with photosynthetic bacteria for CO₂.

Inhibition due to accumulation of hydrogen increasing partial pressure.

Dependence on oxygen for electron transfer.

Lack of enzymes required for VFA breakdown.

Q9. Which type of methanogen performs acetoclastic methanogenesis?

Methanothrix species

All bacteria in Firmicutes

Any organism performing fermentation

Hydrogenotrophic archaea only

Q10. Which condition can strongly inhibit methanogens?

High oxygen levels

Low salinity

Absence of VFAs

Low ammonia concentrations

Q11. Why is syntrophic acetate oxidation (SAO) important when acetoclastic methanogens are absent?

It bypasses hydrolysis entirely.

It converts acetate into CO₂ and hydrogen, enabling methane formation by hydrogenotrophic methanogens.

It lowers pH to stimulate fermentation.

It prevents VFAs from forming in the first place.

Q12. Why are thermophilic digesters less common despite faster reaction rates?

Thermophilic microbes cannot produce methane.

They are more unstable and harder to operate reliably.

They generate only CO₂, not methane.

They require expensive oxygen injection.

Q13. What does Organic Loading Rate (OLR) describe?

The amount of oxygen supplied to the digester daily.

The quantity of organic substrate added per unit volume per day.

The methane concentration in digestate.

The maximum temperature microbes can tolerate.

Q14. Why is methanogenesis considered the 'star of the show' in anaerobic digestion?

It produces acids that stabilize pH.

It is carried out by the largest number of microbial species.

It creates methane, the main energy-rich product.

It requires no specialized enzymes.

Q15. Anaerobic digestion requires fully anaerobic conditions.

True

False

Q16. Biogas is mostly composed of methane and carbon dioxide.

True

False

Q17. Co-digestion is done randomly without considering substrate characteristics.

True

False

Q18. BMP tests can determine both methane potential and degradation speed.

True

False

Q19. Volatile Solids (VS) represent the inorganic fraction of a substrate.

True

False

Q20. Hydrolysis can be accelerated using pretreatments such as grinding or chemical treatment.

True

False

Q21. Acidogenesis is typically the fastest step in anaerobic digestion.

True

False

Q22. High hydrogen partial pressure inhibits acetogenesis.

True

False

Q23. Methanogenesis is performed by both bacteria and archaea.

True

False

Q24. Acetoclastic methanogens convert acetate directly into methane.

True

False

Q25. Thermophilic digesters operate around 50–60°C and have faster reaction rates.

True

False

Q26. A high Organic Loading Rate always increases methane production safely.

True

False

Q27. VFAs accumulate when acidogenesis is much faster than methanogenesis.

True

False

Q28. Hydrogenotrophic methanogens consume hydrogen, maintaining favorable conditions for acetogenesis.

True

False

Q29. Methanothrix is a key genus performing hydrogenotrophic methanogenesis.

True

False