Transcript Novel biotechnological processes for production of polymers

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Utilization of Waste in
Industrial (White) Biotechnology
White Biotechnology
• is an emerging field within modern biotechnology that
serves industry.
• It uses living cells like moulds, yeasts or bacteria, as well
as enzymes to produce goods and services. Living cells
can be used as they are or improved to work as "cell
factories" to produce enzymes for industry.
• White Biotech can help realize substantial gains for both
environment, consumers and industry.
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Biotechnology
Health
Sustainability
Red Green
Health Care Agro-Food
White
Industrial
Using nature’s toolset
Unmet Needs
Economy
Industrial (White) Biotechnology
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Cell factories
Sugars
Biofuels
Biomaterials
Biochemicals
The IB Value Chain
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Bulk
Biofuels
H2
Ethanol
Feedstocks
-Renewable
- Fossil
Sugars
Biomaterials
Polylactic acid
1,3 propane diol
PHAs
Bioprocesses
Biochemicals
Food Ingredients
Pharmaceuticals
Fine Chemicals
Fine
Industrial Biotechnology
Cleaner and more (cost) efficient ways of making:
Faded jeans
Detergents
Plastics
Vitamins
Antibiotics
Fuel
Biosteel
Biobatteries
DNA computers
Reduced environmental
foot-print up to 20 – 60 %
Added Value of
11-22 billion € per Year
IB: three P’s go hand in hand
People
sustainability
Profit
Planet
The IB Value Chain
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Bulk
Biofuels
H2
Ethanol
Feedstocks
- Renewable
- Fossil
Sugars
Biomaterials
Polylactic acid
1,3 propane diol
PHAs
Bioprocesses
Strong points Europe
• Enzymes
• Biochemicals
Biochemicals
Food Ingredients
Pharmaceuticals
Fine Chemicals
Fine
The IB Value Chain
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Bioenergy
Biofuels
Biomass
Feedstocks
-Renewable
- Fossil
Bulk
H2
Ethanol
Sugars
Biomaterials
Polylactic acid
1,3 propane diol
PHAs
Bioprocesses
Biochemicals
Food Ingredients
Pharmaceuticals
Fine Chemicals
B&B
Fine
Developing a Strategic Research Agenda and Roadmap (1)10
Main R&D objectives
Strain, biocatalyst & process optimization
Novel and/or improved functionalities and products
Developing a Strategic Research Agenda and Roadmap (2)11
Research & Technology areas in IB
• Novel enzymes and microorganisms – metagenomics
• Microbial genomics and bioinformatics
• Metabolic engineering and modeling
• Performance proteins and nanocomposite materials
• Biocatalyst function and optimization
• Biocatalytic process design
• Innovative fermentation science
• Innovative down-stream processing
• Integrated biorefineries
Novel biotechnological processes
for production of polymers,
chemicals, and biofuels from waste
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Background
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 Ecological reasons to promote „White Biotechnology“:
Global Warming, Green house effect
 „Rio Declaration on Environment and Development“ June 1992:
Broad consensus to switch to alternative, sustainable Technologies
Principle 4:
„In order to achieve sustainable development, environmental
protection shall constitute an integral part of the development
process and cannot be considered in isolation from it.“
 Rising Prices for mineral oil: Economic necessity to promote
technologies independent from the availability of fossil feedstocks
 Major Drawback for „White Biotechnologie“: Costs for Raw Materials
 Solution Strategy: Utilization of Waste Materials for Production of
Biopolymers, Biochemicals and Biofuels
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Need for „White Biotechnology“ for Production of
Biopolymers, Biofuels and Biochemicals?
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June 2008: Price surmounted 130 US-$ per barrel
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July 2008: Price surmounted 140 US-$ per barrel
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„Target Areas“: Final Products from Conversion
of Waste Materials via „White Biotechnology“
Final Products:
Biopolymers
(PHA, PLA)
Biofuels
(Bioethanol,
Biodiesel)
Biochemicals
(Fine chemicals,
Organic acids,
Antibiotics, Aromatics,
Surfactants, Solvents,
Chiral Synthons)
Catalytically active
Biomass for Production
of Biopolymers, Biofuels
and Biochemicals
Major substrates for production of
biopolymers, biofuels and biochemicals:
Industrial producers
of Waste streams:
Monosaccharides: Glucose,
Galactose, Fructose, Xylose,
Arabinose
Disaccharides: Sucrose,
Lactose, Maltose, Cellobiose
Dairy Industry: Whey
Sugar cane industry: Molasses, Bagasse
Wood processing industry,
Paper Industry
Polysaccharides: Starch,
Cellulose, Lignocellulose
Additional agricultural
branches (e.g. straw from rice,
mais etc., olive oil production, palm
oil industry, sugar beet industry)
Organic acids
Alkohols: Glycerol, Methanol
Biodiesel production: raw
glycerol phase, low-quality biodiesel
fractions
Lipids
Proteinaceous
materials (Peptides)
Slaughterhouses and Rendering
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Industry: Meat- and Bone Meal, slaughter
wastes
Brazil: Integration of Biofuel & Biopolymer
Production into Sugar Cane Industry: Actual and
Potential Utilization of the waste streams
Sugar Cane
2,160.000 t/a
Fibers potential filler for PHA-based materials?!
561.600 t/a
Combustion
Bagasse
Bioethanol
Steam and
32,4 GW/h / a
electrical power
Milling
Hydrolysis
395.000 t steam/ a
Extraction
Convertible Sugars
Raw Juice
Biofuel
Production
180.000 t/a
Saccharose
Selection of production strain!
Hydrolysis
to Glucose and Fructose
PHA
Biopolymer
Production
30.000 t/a
Molasses
Hydrolysis
to peptides and amino
Higher Alcohols
(Butanol, Pentanols)
Destillation
(Glucose, Xylose, Arabinose)
Crystallization
52.575 m3/a
Fermentative
Conversion to Downstream
Bioethanol Processing:
Extraction of PHA
1.) Production of
from biomass
catalytically
active Biomass
2.) Production of
PHA biopolyesters
Residual Biomass
acids
Extraction
solvents!
PHA
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Biopolymers
10.000 t/a
PHB INDUSTRIAL S/A – Sao Paulo,
Brazil
Production of PHB
homopolyester and Poly-3HB-co-3HV copolyesters
from sugar cane saccharose;
autarkic energy supply!
Basic research: TU Graz,
Austria
View of the PHB Pilot Plant for 50 t/a
Production strain: Cupriavidus necator DSM 545 (formerly
Wautersia eutropha)
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Intented industrial scale production of PHA: 10.000 t/a
Whey from Dairy Industry – a versatile Feedstock
for Biotechnology
 Application of Whey lactose (D-gluco-pyranose-4-ß-
D-galactopyranoside) from dairy industry: animal feed,
sweets, food processing, baby food, laxatives,
pharmaceutical matrices
 But: annually 13,500.000 t of Surplus Whey in
Europe (620.000 t lactose)!
 Ecological problem; polluting whey partly disposed
in sea
 2001: EU – project WHEYPOL (G5RD-CT-200100591): application of surplus whey from Italian dairy
industry as substrate for PHA biopolyester production
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From Milk to Whey towards PHA
Biopolyesters
MILK
Pasteurization
Transformation (enzymatic or acidic)
Curd cheese
2.) Production of
1.) Production of
PHA biopolyesters
catalytically
Yield PHA/C-source = 0,33 g/g:
active Biomass ca. 200 000 t/a PHA in EU
from surplus whey possible!!
Full Fat Whey
Skimming
Skimmed Whey
(ca. 13 500 000 t/a in EU surplus!)
Pasterization
Storage
Concentration
Lactose Hydrolysis to
Glucose and Galactose ?!
WHEY CONCENTRATE
Ultrafiltration
Whey
Retentate
α-Lactoglobulin (2 wt.-%),
ß-Lactoglobuline (9 wt.%); Lactose (15 wt.-%)
(depends on production strain)
(ca. 2 700 000 t/a in EU surplus!)
Whey
Permeate
Storage
20 – 21 wt.-% Lactose (81% of the entire lactose from milk)
(ca. 620 000 t/a in EU from surplus whey!)
Desalting ?!
(necessity depends on
production strain)
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Different Routes from Whey Lactose
to Biopolyesters
(Koller et al., 2007)
Whey Lactose
Direct
Application of
Lactose (sufficient
ß-Galactosidase
activity of
production strain)
for production of
Hydrolysis
towards Glucose
and Galactose
for Production of
PHA
Bioconversion 1:
via Lactobacilli
from Lactose to
Lactic Acid
Bioconversion 2:
from Lactate to
PHA
Polylactic acid
(PLA)
(www.igb.frauenhofer.de/
WWW/GF/dt/GFDP 21
Molke.dt.html)
PHA
Conversion to Lactic acid esters
Pyrolysis
Lactones
Unsaturated compounds
(Crotonic acid, 2-Pentenoic acid etc.)
→
Green Solvents
Synthons for chemical synthesis
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Alternative Biotechnological Products
from Whey Lactose
 Bioethanol: Golden Cheese Company, California (19.000 m³
Bioethanol/year) (For Europe: Surplus whey would yield 290.000 m³
Bioethanol/year) (www.ethanolfra.org/pr010201.html)
 Antibiotics: e.g. Bacteriocin Nisin (polycyclic peptide antibiotic from
Lactococcus lactis) against highly pathogenic food-spoiling bacteria
Listeria monocytogenes and Clostridium botulinum (Hickmann, Flores, Monte Alegre, 2001)
 Sophorolipids: Emulsifiers and Surfactants for pharmaceutical,
cosmetic and food industry; chemically: sophorose derivates linked to
hydroxy fatty acids
 Two –step process: Yeast Cryptococcus curvatus cultivated on
whey permeate, accumulates single-cell-oil (SCO) from whey
lactose. SCO is converted in a second step to sophorolipids by
Candida bombicola (Daniel et al., 1999))
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www.lipidlibrary.co.uk/Lipids/rhamno/image006.gif
www.profoodinternational.com
The Increasing Amounts of Biodiesel
 Legislative Situation by the European Commission:
Shares of Biofuels [%]:
 2005: 2%
 2010: 5,75%
 possibly up to 20% until 2020 (8 * 1010 Liter/a in Europe)
 2005: Production of 1,925.000 t in Europe (= 192.500 t
glycerol)
 2008: 2,649.000 t in Europe (= 264.900 t glycerol)
 Austria: 2006 Production of 121.665 t Biodiesel; 2007:
241.381 t (+98%!!!)
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Glycerol Liquid Phase: Waste from the Biofuel
Production for the Production of Biopolymers
e.g. Waste Cooking Oils,
waste animal fats
WASTE LIPIDS
Some lipids: direct
application as feedstock!
MeOH (EtOH)
OH-
1.) Production of
catalytically
active Biomass
2.) Production of
PHA biopolyesters
Yield PHA/C-source = 0,33 g/g:
ca. 88 000 t/a PHA in EU
from surplus GLP possible!!
Transesterification
Biotechnological Production of PHA
Biopolyesters
Mixture Biodiesel
-Glycerolphase
Downstream Processing
Demethanolization
Separation
Degreasing
PHA
Biopolyesters
Residual Biomass
(Proteins, Lipids)
Washing, Dewatering
GLYCEROL LIQUID PHASE
(GLP)
BIODIESEL (RME)
Low-quality biodiesel fractions:
excellent feedstock for PHA
production!
typically 2-4 wt.% of biomass
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Lignocellulosic Feedstocks
 Occurence of lignocellulosic waste:
 wood residues (including sawmill and paper mill discards)
 municipal paper waste
 agricultural residues (including corn stover, rice straw and sugarcane
bagasse)
 special energy crops
 Amounts: non-wood lignocellulosic straw alone is estimated with 2,5*109 t/a
 Composition of Lignocellulose:
Carbohydrates
Cellulose fraction
Monomer: Glucose (Hexose)
+
Lignin (Methoxyphenylpropane)
Hemicellulose fraction
Monomers: Xylose, Arabinose (Pentoses)
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Biotechnological Utilization of
Lignocellulose
 Obstacle: Lignocellulose has evolved to resist
degradation and to confer hydrolytic stability and
structural robustness to the plant cell walls by
crosslinking between the carbohydrates and the
lignin via ester and ether linkages
 Focus of research: UPSTREAM TECHNOLOGY:
Enhanced lignocellulose digestion and the
development of EFFECTIVE ENZYMES for the
degradation of cellulose and hemicellulose into
glucose and pentoses are the prerequisite for an
efficient production of the desired bio-products
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Composition of Different Lignocellulosic Materials
Cellulose
[wt.-%]
Hemicellulose
[wt.-%]
Lignin
[wt.-%]
Corn cobs
42 – 45
33 – 35
10 - 15
Corn stover
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25 - 38
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Wheat straw
33 - 47
22 – 30
13 - 19
Hemp straw
44 - 45
19 - 21
20 - 22
Rice straw
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36
10
Bagasse
40
29
13
Beech wood
46
31
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Fir wood
43
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Poplar wood
50
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Conversion of Lignocellulose to
Value-added Bioproducts
Plant Biomass
High energy
input needed!
Steam Explosion
Alternatives
have to be
developed!
e.g.: Solid State
Fermentation!
Extraction with water
Development
of efficient
hydrolysis
methods
required!!
Hemicellulose
Hydrolysis
(enzymatic or chemical)
Pentoses (Xylose, Arabinose)
Fermentation to Bioethanol
Lignin
Alkaline extraction
Adhesives
Hydrolysis
(enzymatic or chemical)
Cellulose
Energy
Biotechnological
Production
of Biopolyesters
Glucose
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Petschacher Barbara, Diploma Thesis, Graz University of Technology, 2001
Follow-up Products of PHAs: Chiral Synthons
for Organic Synthesis
Chiral synthons: Stereoregular compounds acting as chiral precursors, e.g.
production of pharmaceuticals, pheromons, vitamins, antibiotics, aromatics, perfumes
PHAs: Biobased Polyesters consisting mainly
of optically pure monomers
Chiral center
Hydrolysis leads to a rich source of
bifunctionel, R(-)-configurated hydroxy acids.
Market value
polymer itself!
higher
than
for
the
Classical Hydrolysis:
Isolation of PHA
(Seebach et al., 1992; Seebach
and Züger, 1982)
In-vivo degradation
In-vivo degradation of PHA
by adjusting the enzymatic
systems
involved
in
intracellular PHA metabolism
via the cultivation conditions
(C-source, pH, T); excretion
of metabolites
Highly efficient process!
PHA
Optically pure
monomers
acidic alcoholysis of the
isolated PHA
Process rather complex and highly Solvent-demanding!
App. 130 PHA buliding
blocks reported- broad range
of possible chiral synthons
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(Lee et al., 1999)
Meat- and Bone Meal (MBM) from Slaughterhause
Waste & Rendering Industry – a Precious Nitrogen
Source for Biotechnological Purposes
 Classical Utilization of MBM: Animal Feed
 Problem: The emerge of Bovine Spongiform
Encephalopathy (BSE, „Mad Cow Desease“)
 Peak: Infection of 3500 head of caddle weekly in
Great Britain
 Alternative Utilization: Energy production by
Combustion → low value-creation
 2001: Task Force Graz University of
Technology for Safe Utilization of MBM to
produce value-added products!
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Hydrolysis of Meat- and Bone Meal
 Precondition of Safe Utilization of MBM:
Hydrolysis of MBM to destroy prions
Structure of a Prion
Causing BSE
Hydrolysis time [h]
SDS-Gel-Electrophoresis of alkaline Hydrolysis of MBM30
(PhD thesis José Neto, Graz University of Technology, 2006)
Production of Meat- and Bone Meal
Possible: Removal of Lipids prior to
hydrolysis („Degreasing step“)
Application of lipids for Biodiesel
Production or as carbon source for
fermentative Production of e.g.
Biopolymers
Application of
hydrolyzed MBM for
Biomass production
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Concluding Remarks
 A broad range of waste materials from different origins
exists to be potentially utilized for biotechnological
production of biopolymers, biofuels and biochemicals
 Selection
of the appropriate waste stream for
biotechnological purposes depends on the global region
where the production is intended. Facilities for production
should be integrated into existing production lines,
where the waste streams directly accrue (Prime example:
Integration of sugar-, bioethanol and biopolymer production
in Brazil)
 Improvement of upstream technologies, selection of
optimized
biocatalysts,
enhanced
downstream
processing and autarkic energy supply are required to
achieve cost efficiency in the production of biopolymers,
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biofuels and biochemicals from waste.
Content :
 Limitation and rising prices of fossil feedstocks and the increasing
need for „White Biotechnology“: Ecological and Economic needs
 „Target Areas“: Final products from conversion of waste
materials via „White Biotechnology“
 What waste materials are available for biotechnological purposes
(occurence and the challenges of their utilization)
 Meat and Bone Meal (Slaughterhouses and Rendering
industry)
 Sugar Cane industry – Integration of Biofuel and
Biopolymer Production
 Whey (Dairy Industry)
 Raw Glycerol Liquid Phase (from Biodiesel Production)
 Waste Lipids
 Cellulosic and Lignocellulosic Feedstocks
 Follow-up Products of PHAs: Chiral Synthons for Organic
Synthesis
 Summary
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 THANK YOU
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