Biofuels and industrial

chemicals from microbes

Fossil fuels
Limited supply
Polluting
Used also for chemicals for synthesis of e.g.
plastics
Eventually oil and coal will be too precious to burn
anyway

Biofuels and Industrial

chemicals from microbes
Can be liquid or gas
From renewable biomass, frequently
waste from other processes, no dwindling
stocks
Less polluting
Microbes can also produce chemical
feedstocks for syntheses

Alkanes: methane
Natural gas from gas/oil fields or town
gas produced from coal
Produced by anaerobic bacteria
(methanogens) found in swamps, organic
sediments and rumen of ruminants
Exploited locally as methane digesters
where economic animal faeces
converted to usable fuel

Methane production by
microbes
By a mixed population of bacteria
First population hydrolyse fats, proteins and
carbohydrates to fatty acids, amino acids, and
sugars
Second population metabolize these to
organic acids (acetic [mainly] propionic and
butyric)
Third population oxidize acids to
corresponding alkanes and carbon dioxide
Carbon dioxide, acetic acid converted to
methane

Butanol
Acetone, butanol, butyric acid and
isopropanol (plus higher homologues)
obtained from Clostridium spp.
fermentations
Fermentable base usually starch,
mollasses, or hydrolysed cellulose
Amount of each product depends on strain
and fermentation conditions

Clostridial fermentations
Acetone-butanol (C. acetobutylicum)
yields also butyric, acetic acids, acetoin,
ethanol CO2 & H2
Butanol-isopropanol (C. butylicum) yields
also butyric, acetic acids, CO2 & H2
Butyric-acetic acid (C. butyricum) yields
also CO2 & H2

Industrial acetone-butanol
fermentation
Weizmann - basic research on Clostridial
fermentation
Acetone produced used to make cordite
(explosive) during W W I
Produced riboflavin as a byproduct
Butanol
feedstock for lacquers, rayon, plasticizers, coatings,
detergents, brake fluids, butadiene (synthetic rubber)
solvent for fats, waxes, resins, shellac, varnish
extractant/solvents in food industry

Butanol today

Largely from petroleum in the West

Beer molasses in USSR
Still performed in China
As the price of oil escalates fermented
butanol may become important again

Butanol as a fuel
Fuel properties
Octane enhancing properties when mixed
with gasoline
High heat of combustion -- butanol: water 4:1
equivalent to anhydrous ethanol
More volatile than methanol or ethanol
Highly miscible with gasoline and diesel

Butanol production
Batch process
Unstirred -- gaseous by-products mix
fermentations
Starch or molasses fermentable base (corn
and by products in US)

Butanol fermentation

anaerobiosis achieved with carbon dioxide purge

low level of inoculum
pH drops as organic acids formed
pH rises as acids metabolized to neutral
solvents
final concentration 2 %
acetone/butanol/ethanol 6/3/1
final yield 37%
carbon dioxide by-product sold
products recovered by distillation

Problems with butanol

fermentation

Lactobacillus contamination
Phage infection
Product inhibition
Cost of distillation
Multiple products
Commercial viability depends on oil price

Industrial ethanol
30 billion litres globally per annum
70 % by fermentation
30 % by catalytic hydrogenation of ethylene

Fermentation ethanol
12% for beverages
20% solvent/feedstock
68% fuel

Ethanol as a fuel
Can be used alone on as a mixture
"Gasohol" ethanol mixed with gasoline

Brazil makes 46 % global ethanol -- offsets oil

imports -- but now has to import ethanol!
Brazilian ethanol from cane sugar (sucrose)
fermented by yeast
can use fermentable bases from plant and dairy
waste
complex polysaccharides require pre-processing

The world is going towards biofuels

16

The world is going towards biofuels

17

18

Sugar/Starch feedstock

Sugar cane - sucrose

Brazil

Corn - starch
United States

Not a preferred option for NZ

Fuel alcohol in Brazil

Sugar cane: 53% converted into alcohol 47% converted
into sugar
Sugar: 50% domestic market
Ethanol production capacity: 16.0 billion litres

Ethanol production: 12.5 billion litres or 205,000 barrels

of oil equivalent per day

Total production of sugar & alcohol: 40 million tons of

sugar equivalent.

Ethanol from maize

Dry milling separates oil from starch -- generates
animal feed
Starch gelatinized and digested to hexoses
(mostly glucose) using thermostable amylases
Glucose base fermented by S. cerevisiae In
continuous or batch mode at 32-38 oC and pH
4.5 -5.0
Generate "beer" of about 10% alcohol
Yeast recovered for reuse
Beer distilled, azeotrope distilled (half the cost of
whole procedure)

Improvements to ethanol
fermentations
Use oleyl alcohol as continuous extractant
passed through fermenter
Top layer removed and centrifuged fermentation mix returned to fermenter
Ethanol recovered by flash vaporization less volatile solvent recycled

Alternatives to yeast
Zymomonas spp. afford higher alcohol
yield but have low alcohol tolerance and
ferment only glucose, fructose, sucrose
genetic engineering to improve yield and
range of fermentable substrates
E coli engineered with Zymomonas alcohol
dehydrogenase and pyuvate decarboxylase
produce ethanol under aerobic and anaerobic
conditions

Ethanol from lignocellulosic

material
Billions of tonnes of waste products produced
per year - under -utilized resource
Composition
Lignin (10-35 %) polymer of phenolic alcohols - not
degraded by microbes
Cellulose (15-55 %) polymer of glucose - not readily
degraded by microbes
Hemicellulose (25-85 %) - hexose (glc, galactose,
mannose/pentose (xylose, arabinose) heteropolymer
- few microbes ferment pentoses to alcohol but this is
required for commercial viability

Lignocellulosic feedstock

Plant residues, agricultural wastes, forestry

biomass
Large, cheap resource

Lignocellulose pretreatment
Pretreatment by
milling
chemical pulping
acid/alkali treatment
solvent extraction
steam treatments

Usually require acid (0.5 - 5% hot

sulphuric acid under pressure or conc
hydrochloric acid at ambient temperature)
or amylase treatment for saccharification

Lignocellulose pretreatment (2)

Acids corrosive
Need acid recycling to make process
commercially viable
Acid hydrolysis hard to control
hemicellulose preferentially degraded and resulting
sugars degraded by the acid

Sugars generated: glc, cellobiose

(disaccharide), xylose
Xylose not fermented by yeast
Organisms that ferment xylose produce poor
yields of ethanol, and are ethanol intolerant

Lignocellulose

Crystalline cellulose ~ 44%

Hemicellulosic polysaccharides ~30%
Lignin ~ 26%

Lignocellulose

Pre-treatment

160-260C, steam, acid/alkaline treatment

Cooling required prior to fermentation

Pre-treatment - 160-260C

Saccharification -

Fermentation -

45-50C

25-30C

Contributes to high operating costs of

bioethanol production

Starch To Ethanol

-Reaction Mechanism-

Enzyme+Energy
SUBSTRATE + H2O --------------------------->
PRODUCT
(Starch)
Xtra)
(Dextrins)

Alpha Amylase (SPEZYME Fred,

SPEZYME

Enzyme
SUBSTRATE + H2O ---------------------------> PRODUCT
(Dextrins)

SUBSTRATE
(Glucose)
32

Glucoamylase

(Optidex L400, Fermenzyme)

(Glu

Yeast
----------------------------> PRODUCT
(EtOH,CO2,Biomass)

Conventional Ethanol Production Process

Thermo-Stable
Alpha Amylase

Glucoamylase

Liquefaction

Saccharification

Milo
Corn
Wheat Water
Rye
Barley
Tapioca

GRINDING

33

Fermentation

Alcohol
Recovery
Distillation &
Dehydration

JET COOKER
>100 C
58 MIN
STORAGE
TANK

60 C
810 HRS
(optional)

SLURRY
TANK
SECONDARY
LIQUEFACTION
95 C
~90 MIN

Yeast

pH adjustment steps are not shown

*
DDGS

 Biofuel Technology Innovation

Material production

Preparation/glycation

Selection of high-yield plants

growing in non-cultivatable
land and development of
technologies for growing
such plants

Fermentation

Development of enzymes
featuring higher ethanol
yields and lower costs

Enrichment/dehydration

Development of energy
conservation technology for
separation through
membranes instead of
distillation

Draining

Fermentation of xylose
Use bacterial xylose isomerase to convert xylose
to xylulose (pentose phosphate phosphate
pathway intermediate)
Genetic engineering
make yeast produce bacterial xylose isomerase
themselves
introduce genes from yeasts (Candida, Pichia spp)
able to ferment pentoses (xylose reductase/xylitol
dehydrogenase) into S cerevisiae

Genetically engineered bacteria (Zymomonas,

E. coli) utilize xylose

Fuel ethanol
Main characteristics are in line with
Sustainable Development:
High performance & versatile energy
source
Environmentally friendly & safe
Renewable
Energy efficient

Summary
Possible to produce range of chemicals by
microbial action
Most important is industrial ethanol
Butanol, acetone and organic acids can
be produced by anaerobic fermentations
Commercial viability depends on oil price