lu umwelttechnik sustainability aspects of macronutrients...
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LU Umwelttechnik
Sustainability aspects of macronutrients
production and application
Tanja Wallner
1 Abstract
In this manuscript several mini projects about macronutrients are discussed
according to their production, field of application and according to their environmental
impact. The mini projects range from food industry (Xanthan), to clothes industry
(Cellulose) to biotechnology (Lipases) to bioenergy (Biodiesel). It is also discussed
which aspects of these projects cover the green chemistry principles.
The main findings include biochemical waste treatment and arguments for and
against animal testing, advantages of enzyme usage, cascade utilization, distribution
characteristics, closed reaction conditions, viscose process efficiency, fuel stability
and diesel emission reduction.
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2 Problematic
Sugars fats and proteins are renewable resources with many possibilities for
application.
Renewable resources are to be favored over fossil-based products, but the
production step doesn’t have to be neglected. The step from raw material to
commodity is essential to validate sustainability of the whole system.
Therefor I chose the following parts for my sustainability report about macronutrients:
• Characterization of an industrial scale bacterial produced polysaccharide –
Xanthan
Food
• Spinning viscose fibers from cellulose xanthate; dying of fibers by adsorption
and characterization of obtained fiber
Clothes
• Stability toward oxidation of Biofuels: Preparation and oxidation stability test of
Fatty Acid Methyl Esters (FAME) via Transesterification of Vegetable Oils with
Methanol
Fuel – Energy
• Purification and characterization of membrane proteins
Enzymes
2.1 Questions
2.1.1 Enzymes
Purification and characterization of membrane proteins
Membrane proteins are key player in the human metabolism. Insights into their
structure gives us better knowledge about their function and can be of great help to
treat overweight, diabetes and cancer. In addition to that lipases are also used as
environmentally friendly detergents. They are a very good example to understand
enzymes better. Enzymes make up a vast number of catalysts in various ecological
favorable reactions as for example waste water treatment. In my final concept I will
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try to give a brief introduction info protein engineering, structural characterization of
proteins and try to describe the gained knowledge in respect to science and human
health. I also try to critically evaluate the characterization methods in respect to
sustainability and green chemistry requirements. Maybe I also focus on animal
testing in biology.
2.1.2 Viscose
Spinning viscose fibers from cellulose xanthate; dying of fibers by adsorption and
characterization of obtained fiber
Afterwards we investigate the application of polysaccharides in the clothing industry.
Maybe we could compare commonly used fibers (either renewable (e.g. cotton) and
fossil based (e.g. elastane, polyamide etc.) with fibers from cellulose xanthate in
respect to sustainability
2.1.3 Fuels
Stability toward oxidation of Biofuels: Preparation and oxidation stability test of Fatty
Acid Methyl Esters (FAME) via Transesterification of Vegetable Oils with Methanol
In Addition to that we will synthesize biofuels out of triglycerides and test their
oxidation stability. As the synthesis is carried out on conditions very similar to
industrial production. Hopefully we will establish a greater understanding about
production processes and the problems of fossil fuel replacement. Maybe we could
also compare the stability of commonly used fuels. The substitution and the effects
on the engine and on the environment.
2.1.4 Xanthan
Characterization of an industrial scale bacterial produced polysaccharide – Xanthan
First, we will establish a reasonable data sheet for the poly saccharide Xanthan to
further use it as a bulk chemical and for food industry. Here it is important to get an
impression what green chemistry really is, how the measurements work and what
characteristics are needed to “convert” a raw material into an industrially usable
substance.
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3 State of the Art
3.1 Enzymes: Protein Lipases
Enzymes are sustainable widely used reagents in a vast number of processes. They
speed up reactions by lowering the activation energy to increase the efficiency of the
whole process. The aim of structural biology is to identify and classify unknown
enzyme structures. Although the amino acid sequence of the protein is known, there
is still no reliable method to predict or calculate the 3D folding. The so-called tertiary
structure (3D constellation of the amino acids sequence) can give important insights
into the function of the protein and the human metabolism, as well as strategies in
medical treatment. It enables the possibility to classify proteins and to gain an
improved understanding of biochemistry.
3.1.1 Animal testing
For many biochemical studies animal testing is still a common method.
Recent estimates of worldwide annual laboratory animal use are imprecise and
unsubstantiated, ranging from 28–100 million.i In the following section positive and
negative aspects of animal testing are summarized:
3.1.1.1 Pro Animal Testing
• Animal testing has contributed to many life-saving cures and treatments.
• There is no adequate alternative to testing on a living, whole-body system.
• Animals are appropriate research subjects because they are similar to human
beings in many ways.
• Animals must be used in cases when ethical considerations prevent the use of
human subjects.
• Animals themselves benefit from the results of animal testing. (animal
medicine)
• Animal research is highly regulated, with laws in place to protect animals from
mistreatment.
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• Animals often make better research subjects than human beings because of
their shorter life cycles.
• Animal researchers treat animals humanely, both for the animals' sake and to
ensure reliable test results.
3.1.1.2 Con Animal Testing
• “Animal testing is cruel and inhumane.”
• Alternative testing methods now exist that can replace the need for animals.
• Animals are very different from human beings and therefore make poor test
subjects.
• Animal tests may mislead researchers into ignoring potential cures and
treatments.
• Drugs that pass animal tests are not necessarily safe.
• 95% of animals used in experiments are not protected by the Animal Welfare
Act.
• Animal tests do not reliably predict results in human beings. 94% of drugs that
pass animal tests fail in human clinical trials.
• Humane Society International compared a variety of animal tests with their in
vitro counterparts and found animal tests were more expensive in every
scenario studied.
3.1.1.3 Further Explanations:
“Animal testing is cruel and inhumane.” According to Humane Society International,
animals used in experiments are commonly subjected to force feeding, forced
inhalation, food and water deprivation, prolonged periods of physical restraint, the
infliction of burns and other wounds to study the healing process, the infliction of pain
to study its effects and remedies, and "killing by carbon dioxide asphyxiation, neck-
breaking, decapitation, or other means."
Alternative testing methods now exist that can replace the need for animals.
In vitro testing, such as studying cell cultures in a petri dish, can produce more
relevant results than animal testing because human cells can be used. Microdosing,
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the administering of doses too small to cause adverse reactions, can be used in
human volunteers, whose blood is then analyzed. Artificial human skin, such as the
commercially available products EpiDerm and ThinCert, is made from sheets of
human skin cells grown in test tubes or plastic wells and can produce more useful
results than testing chemicals on animal skin. Microfluidic chips ("organs on a chip"),
which are lined with human cells and recreate the functions of human organs, are in
advanced stages of development. Computer models, such as virtual reconstructions
of human molecular structures, can predict the toxicity of substances without invasive
experiments on animals.
Animals are very different from human beings and therefore make poor test subjects.
Animal tests may mislead researchers into ignoring potential cures and treatments.
Some chemicals that are ineffective on, or harmful to, animals prove valuable when
used by humans. Aspirin, for example, is dangerous for some animal species.ii
3.1.2 Laboratory waste management and Process design
As in many laboratories also in biochemistry laboratories there is a lot of waste
produced. It is necessary to sterilize biological active compounds. In addition to that
radioactive waste from biological tracers must be treated. But most of the time it’s
due to purity reasons that laboratory equipment is used one way. Further
convenience is also a great point. Therefor a thoughtful waste management is
needed. The most important point is to at first raise awareness about laboratory
waste causing a severe impact on the environment. In Addition to that an efficient
working strategy must be developed and passed to successors. As green chemistry
states, “waste prevention” in the first place is crucial and techniques which require
less hazardous substances are preferable.iiiIf the waste is inevitable a good
separation is crucial. Further information iv
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3.1.3 Environmentally friendly and benefiting enzymes
Enzymes are used in industries including agro-food, oil, animal feed, detergent, pulp
and paper, textile, leather, petroleum, and specialty chemical and biochemical
industry. They help to maintain an unpolluted environment through their use in waste
management.v
Examples:
• Waste water treatment, Steroid binding in waste water treatment
• Microbiological biodegradation of petroleum contaminated soil
• Fermentation processes for bioethanol, Cellulases and ligninases for biofuel
• Microbiological enzyme production nucleases, DNA ligase and polymerases
• Xylanases for paper industry
• Proteases and lipases and amylases as detergents, desizining of textile fibers
, The application of cellulases for denim finishing and laccases for
decolorization of textile effluents and textile bleaching
• Acylase racemate cleavage
• Protease cheese production and patain to tenderize meat, Amylases,
betaglucanases, acetolactases for brewing industry, amylase starch
production vi
3.2 Viscose: Textiles
3.2.1 Comparison natural synthetic fibers
Fibers are divided into three main categories:
• Natural – like flax, wool, silk and cotton
• Manufactured – made from cellulose or protein
• Synthetic – made from synthetic chemicals.vii
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The difference between “manufactured” and “synthetic” fibers is that the
manufactured fibers are derived from naturally-occurring cellulose or protein, while
synthetic fibers are not. viii
In 2015 62% of the global fiber consumption was covered by fossil based synthetic
fibers, 25% by cotton, 6% by bio based synthetic fibers 1% by wool and 5% other
natural fibers.
figure 1 global fiber consumption
3.2.1.1 Advantages of synthetic bio-based fibers
Viscose is a man maid manufactured but not synthetic fiber made of chemically
designed cellulose units and therefore called regenerated cellulose fiber. The
advantage of fiber engineering is the possibility to adjust the fiber to our wishes.
Parameters like, thickness, tenacity, elasticity, resistance can be engineered within
the viscose process.
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figure 3 chemical reaction viscose process
3.2.2 Viscose process
In the viscose process, cellulose is treated
with caustic soda (aka: sodium hydroxide) and
carbon disulfide, converting it into a gold,
highly viscous liquid about the color and
consistency of honey. This substance gives its
name to the manufacturing process, called the
viscose process.
The viscous fluid is allowed to age, breaking
down the cellulose structures further to
produce an even slurry, and is then filtered to
remove impurities. Then the mixture is forced
through fine holes, called a spinerette, directly
into a chemical bath where it hardens into fine
strands. When washed and bleached these
strands become rayon yarn.ixx
The Chemical Reaction is shown below xixii
figure 2: viscose process scheme
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3.2.3 Byproducts their environmental impact and toxicity
This process needs sulfuric acid (H2SO4) and Carbon disulfide (CS2) and Hydrogen
sulfide (H2S) is produced as by product. Sulfuric acid, which is used in the spinning
step, is toxic for fish with a LD50 of 1.67 mg and human skin contact leads to burns
and irritation of the respiration when inhalating aerosols.xiiixiv Carbon disulfide is toxic
to organs, influences the reproductivity, irritates the skin and causes a severe damage
to the eyes when exposed.xvHydrogen sulfide, has a LD100 of 500-1000 parts per
million (ppm). Exposure to lower concentrations leads to such as 10-500 ppm, can
cause various respiratory symptoms that range from rhinitis to acute respiratory failure.
H2S may also affect multiple organs, causing temporary or permanent derangements
in the nervous, cardiovascular, renal, hepatic, and hematological systems. xvi
3.2.4 Alternatives: Lyocellverfahren
Pulp are dissolved in N-methylmorpholine N-oxide (NMO). The filtered cellulose
solution is then pumped through spinnerets, devices. The outcoming fibers are drawn
in air to align the cellulose molecules. The fibers are then immersed into a water
bath, where desolvation of the cellulose sets the fiber strands; the bath contains
some dilute amine oxide in a steady state concentration. Then the fibers are washed
with de-mineralised water. The Lyocell fiber next passes to a drying area, where the
water is evaporated from it. NMO exposure leads to irritation of the skin respiratory
system and eyes. Acute toxicity of amino oxide in fish, as indicated by 96h LC50
tests, is in the range of 1,000–3,000 ug L−1 for carbon chain lengths less than C14.
LC50 values for chain lengths greater than C14 range from 600 to 1400 ug L−1.xvii
The amine oxide used to dissolve the cellulose and set the fiber after spinning is
recycled. 98% of the amine oxide is typically recovered. Since there is little waste
product, this process is relatively eco-friendly.xviii
The main advantage is the closed cycle process.
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3.3 Fuels: FAME
3.3.1 Fatty acid methyl esters (FAME)
FAME are a type of fatty acid ester that are derived by transesterification of fats with
methanol. The molecules in biodiesel are primarily FAMEs, usually obtained from
vegetable oils by transesterification. They are used to produce detergents and
biodiesel.
figure 4 transesterification reaction resulting in FAME xix
3.3.2 FAME Production
Transesterification of a vegetable oil is carried out with methanol under catalysis with
potassium hydroxide. For homogenization of the reaction mixture a high-speed stirrer
Ultra-Turrax is used. The reaction is carried out in 2-steps (more steps in industrial
scale), after which the formed glycerol is separated by centrifugation. The obtained
methyl esters are washed with water and dried.
3.3.3 Problems of fossil fuel replacement
Fossil based Diesel is a mixture of hydrocarbons without oxygen in the structure. In
contrast to that bio-based Diesel is a mixture of fatty acid methyl esters with oxygen
in the ester bond. Because of that fact Bio Diesel is less stabile to oxidation than
fossil-based Diesel. Therefore, Oxidation Stability has to be tested to assure a good
driving performance and to be prone of engine damages. The low aromatic and sulfur
content in biodiesel reduces SO2 emissions and particle emissions. In addition to that
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lower hydrocarbon, CO and fine dust emissions are emitted because of the higher
oxygen content.
In contrast to that NOx emissions are higher are observed. These components also
depend on (injection time, ignition delay and adiabatic flame temperature. xx
3.3.4 Emissions/exhaust fumes
The main pollutants are listed below:
• Carbon dioxide (CO2)
• Carbon monoxide (CO)
• Nitrogen oxides (NOx)
• Ammonia (NH3)
• Polycyclic aromatic hydrocarbons
• Volatile organic compounds
• Sulfur dioxide (SO2)
• Particle emissions
• Soot
• Fine dust xxi
3.3.4.1 Catalytic converters
These are the most common catalysts for exhaust fume waste treatment
• Tree-way catalytic converter
• The NOx-Reduction catalyst
• Selective catalytic reduction via urea
• Particle filter
The Tree-way catalytic converter converts carbon monoxide, nitrogen oxide and
hydrocarbons into carbon dioxide, nitrogen and water.
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The reaction steps are:
Reduction of nitrogen oxides to nitrogen
Oxidation of carbon monoxide to carbon dioxide
Oxidation on unburnt hydrocarbons to carbon dioxide and water
figure 5 reaction in the converter xxii
3.4 Xanthan: Food Applications
3.4.1 Xanthan polysaccharide
Xanthan Gum is a 1,4-linked beta-D-glucose polysaccharide with five sugars
containing different residues. It forms helical fibers. The indicated (distribution)
molecular weight of Xanthan is around 2000 kDa.
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figure 6 Xanthan Gum structure
In Aqueous solution it forms gums and gels and is used as flocculant, binder. Film
former, lubricant and friction- reducer. It is industrially produced via fermentation with
the bacterium Xanthomonas campestris. Due to aggregation most of the material
characteristics an only be given in form of distributions. In general, it is highly viscose
also in low concentrations. It shows high viscosity at low shear rates and a high
elastic modulus. It is stable to ionic variations, heat, change in pH, shear stress,
enzymes, and chemicals like salts, acids and bases. It is used in food industry for
texture and freezing stability in baked goods, beverages, confectionery, dairy
products, desserts and dressings. In addition to that it is used in personal care
applications as in dispersed oil phases, shampoos, make-up, lotions and creams,
and liquid soaps. Further it is used in pharmaceutical applications and oral care
applications like toothpastes. In industrial applications it is used in acid-based
cleaners, animal feed, surface coatings, carpet printing and fire-fighting foam xxiii
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3.4.2 Datasheet (relevant numbers)
A useful datasheet has to include all relevant characteristics of the molecule.
In case of a polymer the chemical composure (REACH registered Monomer),
dimensions, geometry and field of application and are needed. In Addition to that also
interactive properties and dynamic properties (mobility, stability, sheer stress
behavior) must be considered.
As we speak of a colloidal system most of the parameters are given as distributions.
For example:
• Mass distribution
• A distribution for the number of molecules
• Volume distribution
• Degree of polymerization distribution
• Viscosity distribution
It is possible to arithmetically calculate the mean values of these distributions, but it
has to be considered that even though they are comparable to mean values of other
substances, they include muss less information than the whole distribution.
Additionally, the relevance of mean values derived from distribution decreases with
increasing width of distributions and rather are zero-approach estimations than
significant information. Range of distribution, percentages of upper and lower limiting
components, dominant mass or molar fractions and distribution of distinct and
separated populations of components or continuous broad distribution provide much
more information about investigated distributions than mean values.xxiv
3.4.3 Green chemistry
Green Chemistry is an attempt to design chemical processes and commercial
products without toxics and waste. It’s a sustainable approach to include energy
conservation, waste reductant and life cycle considerations.
The main principles of green chemistry are:
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1. Prevent waste
2. Design safer chemicals and products
3. Design less hazardous chemicals syntheses
4. Use renewable feedstock chemicals
5. Use catalysts not stoichiometric reagents
6. Avoid chemical derivates
7. Maximize atom economy
8. Use safer solvents and reaction conditions
9. Increase energy efficiency
10. Design chemicals and products to degrade after use
11. Analyze in real time to prevent pollution
12. Minimize the potential of accidents
In addition to that also main principles of green engineering are formulated:
1. Inherent Rather Than Circumstantial
2. Prevention
3. Atom Economy
4. Less Hazardous Chemical Syntheses
5. Designing Safer Chemicals
6. Safer Solvents and Auxiliaries
7. Design for Energy Efficiency
Use of Renewable Feedstocks
8. Reduce Derivatives
9. Catalytic reagents (as selective as possible) are superior to stoichiometric
reagents.
10. Design for Degradation
11. Real-time analysis for Pollution Prevention
12. Inherently Safer Chemistry for Accident Preventionxxv
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3.4.4 Cascade usage
Further it is fundamental to consider a certain kind of process design hierarchy to
make a process as efficient as possible. This hierarchy called cascade utilization.
This approach prefers the application of renewable resources at first on the food
marked, followed by the chemical market and as the last step biomass for energy
production.
4 Materials Methods Results and perspektives
4.1 Enzymes
To solve a protein structure an assessable source of the protein of your interest is
needed. There are two main approaches to isolate proteins either from an already
available organic sample or in the laboratory via microbiological expression.
figure 7 Steps to get a crystal structure
As a next step, the bacterial colony has to be harvested. The purification step starts
afterwards. Different proteins require different purifications techniques. These
techniques (MBP Column's, Ni-NTA Affinity Resin, Strep Tactin to name just a few)
can be implemented with the help of new technologies like cloning and side directed
mutagenesis. If no bacterial expression is used, the harvesting can be neglected but
there are less possibilities to implement the above-mentioned methods. In the case of
the Lipo-Protein-Lipase (LPL) purification Heparin Colum's are chosen, because a
big domain of LPL shows a very high affinity to them. Afterwards, the separation
Get source / microbiological
expression
Isolate and adjust Protein
Crystalize or solve Protein
Measure structural date
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according to the volume of the Lipase in solution is done by Size Exclusion
Chromatography.
a) b) c)
figure 8: purification chromatograms: Chromatography and Electrophoresis Gels
a) First manual heparin purification. This step: washes Casein and Lactoglobulin out of the solution
b) Second heparin purification step
c) Result of the size exclusion purification:
now the Antithrombin 3 cannot be observed anymore
After the purity is checked by spectrometric measurements and Electrophoresis Gels,
a technique to get structural data has to be chosen. For small molecules NMR is
suitable. In case of larger molecules like proteins, solid state NMR is possible. X-Ray
measurements are chosen is this case because they are the most established
technique at that moment. It should be mentioned that this method is relatively
reliable, but it needs a protein crystal to diffract on.
The crystallization was the main challenge on this project. There are attempts to
crystallize LPL since 1970. To crystallize a relatively amorphous protein, a very high
purity is needed. The yield of an 8L Milk purification is approximately 1mg of Protein.
The pure protein was mixed with a vast amount of (approx. 2000) different
standardized crystal screen conditions. Only 0,5% of these conditions showed a
crystal-like behavior. Weekly microscopical examination for the duration of 6 months
showed 5 promising protein crystals were picked and measured on the X-Ray anode.
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Most of them where salt crystals caused by the crystal screen conditions itself. Only
one crystal could be a LPL crystal but it didn‘t diffract.
figure 9: (top down left right) Promising crystal conditions with their position and exact chemical condition
Crystal Screen 29.4. (P- Buffer) A6-6, 0.2 M Magnesium chloride hexahydrate, 0.1 M TRIS hydrochloride pH 8.5,
30% w/v Polyethylene glycol 4,000
Index 29.4 (P- Buffer), D1-37, 25% w/v Polyethylene glycol 1,500
JCSG 12.5, A1-1, 0.2 M Lithium sulfate 0.1 M Sodium acetate 4.5 50 % w/v PEG 400
Diffracting crystal, bMGL-C14 complex initial
After these projects, other experiments had been planned for ssNMR measurements,
Binding Studies and other reagents to stabilize LPL (e.g. GPIHBP1 Peptide Fragment
experiments and other Assays)
4.2 Viscose
In order to design a durable viscose fiber with new features the raw material and
biopolymer cellulose has to be broken down via the viscose process. In this lab we
got a sample of Cellulosexantate from Lenzingxxvi. We made solutions with a different
amount of cellulose nano-crystals and compared the resulting fiber thickness. In
addition to that we also measured the reactant (Cellulosexantate) and the product
(viscose) via IR spectroscopy
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Produced viscose fiber
After the viscose process two monomer glucose subunits contain a maximum of one
xanthate residue. xxvii
figure 10 Cellulose xanthate after the viscose process
figure 11: IR spectra viscose fiber
We tried to wash off all Xanthate residues. Infrared measurements showed that this
washing step hast to be improved to get rid of every single sulfur, indicated by the peak
at 1640 1/cm for double bonds (in our case C=S double bonds).
4.3 Fuels
FAME
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The FAME was produced through esterification of vegetable oils and analytically
characterized as follows:
Ester content:
• FT-Infrared Spectroscopy (FT IR)
• Thin Layer Chromatography (TLC)
• Gas chromatography with flame ionization detection (GC FID)
• Gelpermeation Chromatography (GPC/SEC)
Oxidation stability:
• Petrooxy,
• Ranzimat
Viscosity and density
• Viskosimeter
GC FID Measurements are standardized EN 14214 norms. The ester content has to
be above 96,5 %.
Table 1: results biofuels
Öl Pflanze IV
(Iod-
Zahl)
[g
I2/100g]
Oxidationsstabilität Estergehalt [%]
PetroOxy
® [min]
Ranzima
t [h]
FTIR TLC
(circa
-
Wert)
GC-
FID
GPC
A Olive 75,1 26,68 14,61 87,1
8
90 96,8 85,1
4
B Raps 105,5 26,03 3,20 88,6
9
85 88,4 82,5
5
C Sonnenblum
e
133,7 13,91 2,52 92,1
4
85 99,0 90,9
4
D1 Distel - - - 88,9
8
- - 84,8
5
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D2 Distel 91,78 25,87 1,32 90,9
7
90 95,9 86,5
2
E Maiskeim 119,3 23,83 4,27 91,9
1
85-90 88,1 83,0
7
Table 2results viscosity density
Öl Pflanze
Dichte [kg/m3]
Viskosität
[mm/s2]
Öl
Methylest
er Öl
Methylest
er
A Olive 898,9 868,4
39,40
7 6,829
4.4 Xanthan
Not finished yet
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5 Summary
Enzymes are sustainable widely used reagents in a vast number of processes. They
speed up reactions by lowering the activation energy to increase the efficiency of the
whole process. Structural Biology gives insights into the mechanism of this molecular
machines and helps to gain a better understanding and more efficient biological
processes. As in every laboratory also in biochemistry labs many different kind of
wastes are produced and depending on the scientific topic also animal testing can be
include.
Xanthan is produced microbiological with help of enzymes. This polysaccharide is
used according to cascade process principle. (Food before Chemicals - Chemicals
before Bioenergy). In addition to that renewable resources and non-hazardous
substances are used during the production but it is very difficult to get feasible
characteristics of the product.
As a green chemistry principle (Minimize the potential of accidents) cellulose
production is performed in a closed circle to prevent emissions but there are still
improvements to make in the field of waste treatment.
As the last step of cascade utilization energy can be processed from biomass. In this
case fuels with a low iodine value are more stable than fuels with a high iodine value.
In addition to that we could use waste from agricultural overproduction, biorefineries
or butchery waste. Further it should be mentioned that biodiesel causes less
emissions than fossil-based Diesel. It would be possible to add more Biodiesel
(nowadays 7%) to the daily used engine fuel.
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i http://www.animalexperiments.info/studies/numbers/worldwide.html
ii https://animal-testing.procon.org/
iii https://www.acs.org/content/acs/en/greenchemistry/what-is-green-
chemistry/principles/12-principles-of-green-chemistry.html
iv https://www.ncbi.nlm.nih.gov/books/NBK55885/
v https://www.ncbi.nlm.nih.gov/pubmed/15493529
vi https://de.wikipedia.org/wiki/Enzym#Enzyme_in_der_Technik
vii http://www.coatsindustrial.com/en/information-hub/apparel-expertise/know-about-
textile-fibres#fibre%20classification 27.05.2018
viii https://oecotextiles.wordpress.com/category/fibers/viscose/
ix https://oecotextiles.wordpress.com/tag/lyocell/
x https://oecotextiles.wordpress.com/category/fibers/viscose/
xi http://costfp1205.com/wp-content/uploads/2017/04/13.-
Roder_final_COST_Stockholm_March_2017.pdf
xii https://de.wikipedia.org/w/index.php?title=Viskosefaser&oldid=177831417.
xiii https://echa.europa.eu/de/registration-dossier/-/registered-dossier/8699/6/2/2.
xiv https://en.wikipedia.org/wiki/Sulfuric_acid#Industrial_hazards
xv www.seilnacht.com/Chemie/ch_cs2.html
xvi https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2850187/
xvii https://en.wikipedia.org/wiki/Amine_oxide#Environmental_safety
xviii https://de.wikipedia.org/wiki/Lyocell#Herstellung
xix https://en.wikipedia.org/wiki/Fatty_acid_methyl_ester
xx https://de.wikipedia.org/wiki/Biodiesel#Abgasemissionen
xxihttps://www.ivt.tugraz.at/images/stories/Files/Skripten/Folien_Sams_Schadstoffbild
ung_Teil_1_2016.pdf
xxii https://secure.lambdapower.co.uk/TechNotes/Tech-10.asp
xxiii https://www.cpkelco.com/products/xanthan-gum/ visited 2018.05.24
xxiv http://ahgroup.at/user_files/SE-&-VO/lecture07/polymerCharacteristics.php
xxv https://www.acs.org/content/acs/en/greenchemistry/what-is-green-
chemistry/principles/12-principles-of-green-chemistry.html
xxvi https://www.lenzing.com/produkte/ 22.05.2018
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xxvii http://kirste.userpage.fuberlin.de/chemistry/kunststoffe/viskose.htm. (Accessed:
2nd June 2018)