Some good news about potentially expanding ethanol use.

EPA Makes Progress Toward Approval of E15

ACE Urges Agency to Approve E15 and Higher for All Passenger Vehicles

Inhibitions from oligomers

An intersting research paper with new findings: "xylobiose and higher xylooligomers were shown to inhibit enzymatic hydrolysis of pure glucan, pure xylan, and pretreated corn stover."

"Thus, b-xylosidase alone does not appear to be sufficient to hydrolyze high DP soluble xylooligomers, and supplementation with both xylanase and b-xylosidase appears desirable to realize high monomeric xylose yields, as found for pure xylan and AFEX and SO2 pretreated corn stover."

What about other oligomers such as mannooligomers in softwood?

Pretreatment: Low temperature and atmospheric

Here is a paper on low temperature pretreatment. What other methods?

• Alkaline peroxide?
• Oxygen-alkaline treatment?
• Ozone?
• Liquid ammonium?

All of these work but which one is more practical in terms of environmental issue and cost?

What about biopulping process?

Two new papers on genetic modification of biofuel feedstocks

1. Genetic modification of lignin biosynthesis for improved biofuel production

2. Genetic improvement of C4 grasses as cellulosic biofuel feedstocks

Plant genetic engineering for biofuel production- a problem solver from the root

"Genetically engineering plants to produce cellulases and hemicellulases, and to reduce the need for pretreatment processes through lignin modification, are promising paths to solving this problem, together with other strategies, such as increasing plant polysaccharide content and overall biomass." A review paper published in Nature summarized the research progress in these aspects.

Cheap sugar: How can it be cheaper?

It is known that feedstock cost account for significant part of overall ethanol production cost. The key is to obtain cheap fermentable monomer sugars from lignocellulosic biomass with the following factors:
· Monomer sugar recovery yield. If we look at biomass, only ~65-70% of cell wall is carbohydrate that is our target to convert into monomer sugars. The question is how to efficiently hydrolyze it with high yield. Chemically or enzymatically? Currently both cannot achieve high sugar yield and high efficiency. The combination of chemical and enzymatical hydrolysis is the direction. However, more factors need to be considered to achieve high sugar yield when talking about a commercialized technology:
o Recoverable/recyclable chemicals that minimize the chemical cost and waste treatment
o The pH of pretreatment that impacts reactor metallurgy
o The temperature of pretreatment that impacts energy usage
o The robust of enzymes that reduce the loading and increase the efficiency

Effects of Cellulose Crystallinity, Hemicellulose, and Lignin on the Enzymatic Hydrolysis

It is known that the efficiency of enzymatic hydrolysis was affected by the degree of cellulose crystallinity, hemicellulose and lignin removal. A recent published paper reported the results on how these factors impacted enzymatic hydrolysis.

Prefermentation: an approach to improve co-fermentation in SSF

Prefermentation was reported as a possible means to overcome the problem of competitive
inhibition fron glucose on xylose: i.e. the free hexoses initially present in the slurry, were fermented before adding enzymes.

Silica bodies: possible physical barriers to chemical penetration during pretreatment

It is known silica content is high for non-woody lignocellulosic biomass.The deposition of silica bodies in biologically engineered craters was found to be a unique feature, which leads to some silica-rich spots on the surface of cell wall. On one hand, the silica will cause scaling problems if not treated properly; on the other hand, they also become barriers for chemical pretreatment. It is neccessary to dissolve the silica through pretreatment. Therefore, the chemicals used and the pH for pretreatment need to be chosen carefully.

A new approach: single step bioprocessing

Low Risk Biorefinery: more product streams

Bioethanol production often faces survival challenging due to its lower margin profit at low oil price and downturn economic recession, which always puts a sole-ethanol production business in a high risky situation. Although there is an incentive from government support with policy, funding, and tax credit, whether it is sustainable profitable will determine the future of this industry. The relatively safe way is to have multiple bio-product streams that are composed of high value-added biochemicals such as lactic acid, succinic acid, glucaric acid to reduce the risks.

US-DOE top 12 building blocks give us the direction.

Partial recycling of pre- treated biomass will increase C6 sugar recovery yield

Paper mill sludge: a cheap sugar for biofuel/biochemicals

Modern paper companies produce large quantities of sludge when using recycled fiber. These sludges are the residue leftover from the paper recycling process and consist of unusable short cellulose fibers inks and dyes, clay, glues and other residue along with any chemicals used in the recovery process. The main disposal routes for paper sludge are land-spreading as agricultural fertilizer or incineration in CHP plants at the paper mill. However, the shortage of landfill space and more restrictive environmental regulations have made disposal more costly and less desirable.

A recent analysis of the sludges from a carton paperboard mill and a tissue paper mill indicating the cellulose are composed of 50-65% of total dry weight of sludge, which can be an important fiber source for biofuel and biochemical feedstock. These fibers with low amount of lignin make them more digestible for enzyme and the process is much simpler compared with the one from biomass. Prior to hydrolysis, there is a need for pre-cleaning or separation of the contaminants through mechanical/chemical process.

The chemical kinetics of the sulfuric acid hydrolysis

The chemical kinetics of the sulfuric acid hydrolysis of softwood was determined by J.F.Seaman in 1945 as follows:

• a 100% increase in acid concentration causes an increase of 153% in the k1 cellulose-hydrolysis-rate kinetics constant, but that increase only caused the k2-sugar product-degradation kinetics constant to increase 103%.
• a 10 degree rise in C temperature causes an increase of approximately 190% in the k1- cellulose-hydrolysis-rate kinetics constant, but then k2-sugar-product-degredation kinetics constant increased only 130%.

Corn steep liquor: a sole cheap nutrient for fermentation

In the lab, expensive yeast and peptone extracts have been used to provide these nutrients for fermentation. However, the cost of the fermentation medium is also one of the principal factors that determines the economic viability of the ethanol production, it is very necessary to find low cost medium components that can supply all the nutritional requirements for good growth and fermentation activity.

Corn steep liquor (CSL) is a liquid by-product of wet milling process of maize-starch industry.It is a sole inexpensive source of nitrogen, vitamins, amino acids and other nutrients, which has been used in the lab study of fermentation and demonstrated the favorable effects compared with traditional complex media formulations for the fermentation.

Thar Process may bring a change on bioethanol recovery

Currently, bioethanol producers are facing another challenging time in its history for viability due to the bad economic situation, volatile commodity markets, and high feedstock cost (if use corns). Some corn-based ethanol producers have been forced to file for bankruptcy before any cellulosic ethanol commercialization takes off.

The key to change the fate of corn-cellulosic ethanol production is still upon the product cost. Cheap feedstock is the first priority. However, any new ideas or technologies that can drop current production cost will be expected in this industry.

Recently, Thar Process receives grant from the state of Pennsylvania for distillation replacement technology, i.e.use a high-pressure propane extraction to remove the high volumes of water from fermented broth and recover bioethanol to replace an existing ethanol plant’s conventional distillation. The propane used in the extraction process can be recycled; a significant energy savings is expected to save from bypassing the conventional distillation and molecular sieve drying steps.

We are looking forward to seeing the progress and economic evaluation of the process.

Recirculation of process streams from fermentation

When SSF fermentation is conducted via yeast, it is suggested to recirculate the process stream. The purpose is to maintain a high concentration of ethanol and dissolved solids and therefore to reduce the energy requirements in distillation and evaporation units ( expensive steps) and reduce fresh water usage. Based on the study by Alkasrawi et al (2002) for SSF process,when 60% of the fresh water was replaced by stillage, ther was no changed for ethanol yield and productivity but ethanol production cost reduced by 17%. When the liquid after SSF was partially recycled, 40% fresh water was replaced;final ethanol yield was not affected with initial productivity decreased; ethanol cost reduced 12%.

However, the disadvantage is that ccumulation of compounds released, or formed in the hydrolysis and fermentation steps may be toxic to enzymes or yeast.

So, we at least have effective enzymes and robust microorganisms for fermentation for this initial consideration of stream liquor circulation, right?

Acetyl group in xylan: problem or opportunity?

It is known that ~7 acetyl groups per 10 xylose units in heardwood and straws, which leads to the formation of acetic acid or acetate by peeling off during any thermal and chemical reaction.

It is quite a big amount in the pretreated hydrolyzate, which is toxic to microorganism at certain level. It is not economical by just removing it from the hydrolyzate. It should be recovered or utilized. Distillation is not efficient to recover it from the hydrolyzate. Membrane separation can achieve the goal but the capital and operating cost will kill the biorefinery if ethanol is the only product.

One of the approach is to produce ethanol indirectly, i.e first ferment xylose to acetic acid followed by esterification and hydrogenation. As a result, 2 unit of xylose can produce 10 unit ethanol with 10 unit hydrogen. The acetic acid can be used completely.

The questions are:

1. The yield and efficiency of fermentation to acetic acid

2. The cost of hydrogen

3. Cost of hydrogenation

4. more...

What are other alternatives to revover or remove acetic acid?

Cheap sugar: the key for bioethanol to survive

Bioethanol industry is facing another winter time in its history due to current cheap oil/gas price. Most people believe the price of oil will back up again sooner or later. The question is when? The good news is the incentive policy and stimulus fund from the new government that will bring the spring this industry. However, the long term survival will depend on its own economic viability. The key is the cheap sugar and apparently the renewable source is lignocellulosic biomass. Generally only 2/3 of biomass weight is carbohydrate that can be converted into monomeric sugars for fermentation. The question is how to obtain them with a high yield in a cheap way. The current hydrolysis technology is still not good enough to realize this.

Enzymatic hydrolysis is a direction for bioethanol production from lignocellulosic biomass. Ideally a or a combination of chemicals are used to remove both hemicelluloses and delignification simultaneously, the resulting solid is mostly cellulose with more exposed and accessible surface and pores, free chain ends, leading to lower enzyme loading, high enzyme selectivity, and fast rate of hydrolysis.

The cheap sugar is calling on advanced enzyme!

Integration may lower the overall hydrolysis cost

Before the cost of enzyme is down to an economical level, it may be not enough to increase the efficiency and rate of enzymatic hydrolysis of biomass just via a pretreatment.

Because of the nature of hemicelluloses (branched, amorphous, and variety), the severity of pretreatment should be low to avoid hemicelluloses degradation. However the mild pretreatment will not damage cell wall enough and remove part of recalcitrant lignin (the physical barrier and competitive sites for enzyme adsorption). As a result, the efficiency and rate of enzymatic hydrolysis cannot reach the level of what we expect. Ideally, the process should be integrated with several units: pretreatment without washing to pull out hemicelluloses; followed by delignification to remove lignin; enzymatic hydrolysis of delignified biomass with very limited dosage of enzyme to achieve target and high sugar recovery yield. The additional unit may increase capital cost. Considering the saving of enzyme dosage and time, the overall operation and material cost may be lower. If increasing the rate of hydrolysis, the size of equipment can be smaller. Therefore, the integration of process may lead to a overall efficiency.

We plant trees is to have a forest!

Trash Today, Ethanol Tomorrow

Major Advance In Biofuel Technology reported by ScienceDaily.

Ethanol removal from reaction–separation integration

The reaction–separation integration is an attractive alternative for the intensification of ethanol fermentation processes. When ethanol is removed in-situ, the product inhibition will be reduced.The methods to remove ethanol from fermented broth are listed as follows:

· Vacuum extraction, which can be conducted by coupling of fermentor vessel with a vacuum chamber extracting the more volatile ethanol from fermentation broth which allows the partial product removal and the increase of overall process productivity.
· Gas stripping to increase the concentration of sugars in the stream feeding the fermentor and improvement of improves liquid circulation and mass transfer.
· Membrane separation. For example, ceramic membranes can be used to filter cell biomass and remove ethanol during the fermentation.The removed ethanol is then distilled and the resulted bottoms are recycled to the culture broth resulting in a drastic reduction of generated wastewater. The coupling of fermentation with the pervaporation is another case to remove produced ethanol and reduce the natural inhibition of the cell growth caused by high concentrations of ethanol product.

In membrane distillation, aqueous solution is heated for vapor formation, which go through a hydrophobic porous membrane favoring the pass of vapors of ethanol (that is more volatile) related to the vapors of water. The process driving force is the gradient of partial pressures mainly caused by the difference of temperatures across the membrane.
Liquid extraction is to use an extractive biocompatible agent (solvent) that favors the migration of ethanol to solvent phase, a process known as extractive fermentation.

Hopefully,we will have lab or trial date rather than just modeling for the evaluation, right?

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Synergistic effect of toxic chemical compounds in hydrolyzate

The typical inhibitory compounds from lignocellulosic biomass dilute acid pretreatment are sugar degraded products (furfural, HMF, formic acid, and levulinic acid etc), lignin degraded products (vanillin etc), extractives, and some metals.

The maximum concentration of each inhibitor that a microorganism can tolerate varies depends on the factors of types of microorganisms, its adaption to the medium, process, the numbers of inhibitors, and very importantly, their synergistic effect. For example, ethanol production was stimulated by acetic acid only (up to 10 g/l) in medium without furfural, or by furfural only (up to 2 g/l) in medium without acetic acid. However, the combination of both will negatively affect the cell growth, cell mass yield, and ethanol yield, i.e. the toxicity of biomass hemicelluloses hydrolyzates results from the aggregation of several toxic compounds (alcohols, aldehydes, and acid) rather than individual compounds.

Effect of detoxification of dilute-acid pretreatment hydrolyzate

Before detoxification

Detoxification-1

Detoxification-2

After detoxification, almost no sugar loss but significantly remove furfural (HMF) and simple phenolic compounds.

The factors on enzyme transport

Due to the different morphology of cellulase, the rate of transport will be regulated by the following factors:
1. The substrate pore size and shape: the specific surface area accessible to the protein (SSAP)
2. product concentration
3. Substrate adsorption preference
4. Physical barriers

Silica, a problematic metal in of herbaceous biomass

It is known that about 3–10% of total feedstock (dry matter) is the residue remaining after ignition (dry oxidation at 575 ± 25°C) of herbaceous biomass. It is composed of minerals such as silicon, aluminum, calcium, magnesium, potassium, and sodium.

During hot water and acid pretreatment, silica will be extracted and soluble in acid solution. When raising pH using alkali, silica (or silica oxide) will be hydrated to form some kind of flocs. As a result, it will precipitate into cells to cause problems for cell growth and scaling problem for the equipment. It can also form complex with some organic compounds which interfere with the process and even fermentation.

The effect of available cellulose chain ends on the rate of enzymatic hydrolysis

It has been found that available cellulose chain ends directly relate to the rate of enzymatic hydrolysis of biomass, which is controlled by the amount of amorphous regions, pretreatment, and the action of endoglucanases. If a pretreatment just increases the available cellulose surface areas without significant change of cellulose crstallinity, it is possible that the rate usually does not show significant increase. If there is enough cellulose chain ends or not generated fast enough, the increase in enzyme loading often does not show the hydrolysis rate increase.

The best biomass pretreatment technology

The best biomass pretreatment technology is no pretreatment.

Chemical effects enhanced by charging N2 or CO2 during pretreatment

During biomass pretreatment, introduction of N2 or CO2 into the reactor will enhance the chemical changes of cell wall components due to the mechanical effect.

Uronic acids and metals in biomass

Uronic acids and metals in biomass will cause some problems in acid hydrolyzates.

Don't panic when your mass great than 100% for biomass composition analysis

When we do biomass chemical composition analysis, it often turns out the mass add-up of all the chemical components great than or less than 100%, which is very common.
1. Different chemical components are analyzed by different methods and instrument, which cause the errors
2. For sugar analysis, each sugar has different optimal post-hydrolysis conditions, which cause both under and over hydrolysis of the sugars
3. Klason lignin includes some ash and proteins, which are accounted for twice
4. Uronic acid and acid lignin should be included when doing mass balance.
5. The standard solution composition should be as close as possible the real samples

Mechanism of furfural inhibition

• Furfural is metabolized by S. cerevisae under aerobic, oxygen-limited, and anareobic conditions to furfural alcohol.
  –Furfural reduce the specific growth rate, the cell-mass yield on ATP, the volumetric, and specific ethanol production
  –NADH-dependent yeast alcohol dehydrogenase (ADH) is believe to be responsible for furfural reduction.
  –Under anaerobic conditions, glycerol is normally produced to regenerate excess NADH formed in biosynthesis. Glycerol production reduction during furfural reduction suggests that furfural reduction regenerates NAD+.
  –Elevated concentrations of acetaldehyde excreted in the beginning of the fermentation, which was suggested to be due to a decreased NADH concentration in the cell during furfural reduction.
  –Furfural inhibition of glycolytic enzymes in vitro and the direct inhibition of ADH might have contributed to acetaldehyde excretion.
  –Intracellular acetaldehyde acumination suggested to be the reason for lag-phase in growth in the presence of furfural.
  –The model
  »i) furfural reduction to furfural alcohol by NADH dependent dehydrogenases had a higher priority than reduction of dihydroxyacetone phosphate to glycerol
  »ii) furfural caused inactivation of cell replication.

Mechanism of acidic acid inhibition

Uncoupling and intracellular anion acumination
–The drop in intracellular pH resulting from inflow of weak acids is neutralized by the action of plasma membrane ATPase, which pumps protons out of the cell at the expense of ATP hydrolysis. At high acid concentration, the proton pumping capacity of the cell is exhausted, resulting depletion of the ATP content, dissipation of the proton motive force, and acidification of the cytoplasm.
–However, the anionic forms of acetic, formic acid, are lipophobic and do not traverse the plasma membrane in both dissociated and undissociated form, causing a high rate of proton impart. The extent of intracellular anion accumination will be a function of the pH gradient over the plasma membrane.

The Effect of Lignin on Enzyme Hydrolysis and Fermentation

• Barrier to cellulase enzymes
  – limit the efficacy of hydrolysis
• Adsorb cellulases
  –Increase enzyme loading and therefore the cost
• Phenloc compounds partition into biological membrances cause loss of integrity, therefore affecting their ability to serve selective barriers and enzyme matrices.

Less heavily substituted phenolics are the most inhibitory compounds
–Phenols
–Vanillin
–Hydroxylbenzaldehyde

Pulp Mill: A Natural Home for the Forest Product Biorefinery

It is costly to start a greenfield biorefinery plant. When we closely look at pulp and paper industry, it will not hard to find:

• this industry is the largest handler of forest residues (lignocellusics)
• $Millions investment has been completed in infrastructure, which will allow to save ~ 35% of capital cost versus a greenfield plant and utilize the boiler house, generators, control rooms, pipe bridges, water and effluent stations, warehouses, woodyards, wood procurement, storage tanks etc
• Hundreds of highly trained technical professionals are available for cellulosic biorefineries
• Co-production and process integration reduce allocated production cost for both pulp mill and biorefinery plant
• Bioethanol adds additional revenue to pulp and paper mills

There, when we borrow the ideas from paper industry to treat biomass , somehow we should not turn our attention away from this industry. More we can take advantge of them.

Purification of Crude Glycerol for Omega-3 Fatty Acids Production

According to Biodiesel Magazine, a technology was developed by Wen at Virginia Tech that uses glycerin to aid in the production of algae that produce omega-3 fatty acids. For this purpose,the soap and methanol present in crude glycerin must be removed before it can be used to aid in the algae growth.

When adjusting pH of the medium containing crude glycerol for algae to grow, the soap contained in the glycerol will be precipitated out of the medium, i.e. the process used to adjust the pH of the solution causes the soap to solidify and settle out of the glycerin.

Next when the glycerin is sterilized at 120 C before used to cultivate algae, the residual methanol will be evaporated and collected.

Biofuels or biochemicals: Are you ready to get in? (2)

According to the report from Ethanol Producer Magazine, the cellulosic ethanol project with $30 million DOE fund was suspended by Lignol and Suncor. The reasons disclosed are instable energy prices, capital market uncertainty and general market malaise. However, although the technology was not listed as a reason, I guess at least a cost-effective well-developed technology is not ready yet since Suncor will continue to monitor the progress of Lignol's technology.

I feel that it is very wise for both partners to make such a decision at an early stage in the project’s development and no significant costs have been incurred. They are serious about cellulosic biorefining.

My question is: what will happen if they continue to proceed the project with the government funding, or with misleading information? What about other players?

These days, biorefining has been driving by a lot of aggressive and ambitious objectives. The good news is that it can attract more attention and investment;on the other side, technology can not be well developed overnight. A gap often occurs between the ambitous goals and the timeline of available technology development. As a result, if unrealistic goals have been set with significant investment, the risk is not far or near around no matter how we are aggressive or smart or very supportive. In the process, realistic and objective evalution must be provided.

It is great lesson for us to learn from this wise decision.

Biofuels or biochemicals: Are you ready to get in? (1)

According to AE report, although many alternative energy investors suffered huge losses in 2008 when several prominent stocks fell by 70% or more, they have not lost hope in alternative energy stocks and their future. The measured optimism from financial community is also a stimulus to alternative energy under the current deeply challenging business climate. So does the government support!

For biofeuls and biochemicals industry, it is still uncertain when the great potential become reality. To use biomass as feedstocks, the utilization of hemicelluloses is the key because it accounts 20-28% of the chemical components in biomass and will have significant impact on the production cost. However the available cost effective technology for commercialization is not ready yet: the scalable microorganisms to ferment C5 sugars under toxic environment, the process to achieve high sugar recovery yield from biomass….

So it is the time for scientist and engineers to get in. It is also great time for investors to jump in if the opportunity has a solid fundamental support. Certainly, a detailed and solid technical evaluation before jumping in is very critical.

A recommendation for biomass pretreatment reactor

The most widely used lab pressurized reactors for biomass pretreatment are autoclaves, steam guns, etc, which is hard to scale up.

A reactor like M&K lab digester has been widely used for lab study on pulping. The rapid liquor circulation provides excellent chemical/biomass interactions or mixing. The two-vessel design allow us to use steam for rapid heat-up and the cooling pipe line allows us to cool down fast after treatment. Such a kind of design has been scaled up for pulp and paper industry and should be easily done for the biorefinery.

Extractives: Easily negligible chemical compounds from detoxification

When talking about detoxification of biomass hydrolyzate, almost all the research and studies focus on these compounds such acetic acid, sugar degraded products (furfural, HMF etc). However, small amount of extractives exist in the hydrolyzate, which is often ignored. Actually, some of these extractives, naturally exist in biomass for coloration and as insectcides, which are also toxic to miroorganisms. For example, stilbenes are generally very toxic! So, it is not enough to remove just furfural and acetic acid.

Typical extractives in tree:

- Resin acids (no or little in hardwood)
- Fatty acids
- Monoterpenes (turpentine)
- Phenolics (Gallic acid, vanillin, Stilbenes, Flavonoids, Lignans)

- Others (Alkanes, Proteins, Monosaccharides and derivatives)

Extractive components are “small” molecules that can be extracted with a solvent from wood, bark, or foliage.
-Generally, extractives are present in small amounts.
-Extractives vary tremendously within species, between species, and within trees.
-There are thousands of different extractives present in wood.

Enzyme torwards breakdown lingin in biomass hydrolysis

The traditional enzymatic hydrolysis of biomass is to hydrolyze pretreated biomass to release monomer sugars for further processing. A new different approach has been reported recently on the research at MSU: it focused the white-rot fungus that is often found on rotting wood and used for biopulping. The fungus contains the peroxidase enzyme that initiates lignin breakdown. After isolation from the fungus, the enzyme-producing gene was reproduced by introducing it into E. coli. The idea was to to isolate the gene, slice out the DNA and basically have the bacteria eat the lignin.

Efficient ethanol fuel cells could be practical soon

A new catalyst cable of breaking ethanol's bonds and generating electricity could soon result in practical portable fuel cells powered by ethanol, according to Technology Review.

"NEW KIDS ON THE BLOCK"

According to ICSI.com, "There is currently a limited talent pool of experienced sustainability executives...".

"The chemical industry, however, is a different animal when it comes to looking for CSOs. Aside from the required tree hugger and company-conscience roles, sustainability leaders of chemical companies need to be technically and financially savvy, understand how the industry operates from top to bottom, and be a long-term strategic thinker."

Actually, there is a great need of real professionals who has solid technical background and experience in these areas. However, we can not build up the Rome city overnight. It takes time and need more patience... Otherwise, the fake experts will cause more problem than the fresh learners.

Detoxification of hydrolyzate by activated carbon

Activated carbon is known to have high affinities for colored impurities in hydrolyzate of sugar mixtures, and its unit cost is relatively lower and it can be reactivated and reused.

It has been found that the affinities of acetic acid and furfural (and HMF) are much higher than that of the sugars. The key parameters include activated carbon particle size (surface areas), the ratio of activated carbon to hydrolyzate, temperature, and adsorption time, etc.

The activated carbon can be packed into a column and the hydrolyzate will be pumped into the column. The residence time will be determined by the flow rate and the length of the column.

A inline IR or UV detector can be used to monitor the compounds in the eluent.

Integration of pretreatment and enzymatic hydrolysis of biomass

This a new concept: The biomass is first pretreated with water only or with other chemicals. After pretreatment, the treated biomass and slurry is enzymatically hydrolyzed without separating the solid and liquid.

Here is one of the research projects conducted at The BioEnergy Science Center.

Membrane operational modes

Based on operational models, membrane can be devided into two type:

1. Dead-end: The fluid flows at right angle to the membrane

-Deposited particles form “Cake Layer”
-Suitable for more concentrated Suspension

-Need more frequent cleaning

2. Cross-flow: The fluid runs parallel to the membrane

-High shear force near the membrane surface
-Minimize cake formation
-Recycle feed stream
-Require more energy
-Stable flux

Membrane separation for biorefinery

1. Detoxification

Nanofiltration (NF) and Reverse Osmosis (RO) membranes can be use to concentrate sugar and remove toxic compounds such as acetic acids, and sugar degraded products in the hydrolyzate.

2. Downstream processing and purification
Microfiltration (MF) can be used to remove particls (proteins etc) from fermented broth.

3. Desalination
ElectroDialysis (ED) can be used to remove inorganic salts

Membrane Process

The number of downstreaming steps for biobased chemical production strongly influences the quality and the price of the product. The total production costs are determined mainly by the downstreaming rather than by production of the product using fermentation.

After fermentation, a pretreatment of the fermentation broth is required to separate biomass, proteins, and cells. The multivalent salts also need to be removed for purification. he separation of inorganic salts and proteins/cells presents a special problem with the production of valuable substances from renewable raw materials. Therefore, the methods used for downstream processing will play a very important role.

Membrane Process is essentially a separation process based on molecular properties. The advanteges of membrane process include:

• It reduces the number of unit processes in treatment systems


• Potential for process automation and plant compactness


• Much smaller foot print than the conventional plants of the same capacity


• Easy scale-up, expansion and retrofication


• Less or no chemical use and provides highest quality water


• No formation of secondary chemical by-products


• Less sludge production


• Water reuse and recycling

Ceretainly, the disadvantges exist, includng membrane fouling, low membrane life time, low selectivity, and high capital and operating cost.

Based on the driving forces, the following processes are defined as:

Pressure driven membrane processes
Microfiltration (MF)

-Simple screening mechanism
Pore size 0.01 μm - 10 μm
DP » 0.01 to 0.5 MPa
-Low pressure process
-Most effectively remove particles and microorganisms (bacteria)
-High flux
-Colloids/Macromole ---> theoretically pass through the membrane

Ultrafiltration (UF)

-Screening and Adsorption
Pore size 1 - 100 nm
DP 0.1 to 1 MPa
-Membrane is classified in terms of Molecular Weight-Cut off (MWCO) : 1000 - 100,000
-Two layers: a thin (0.1 to 0.5 µm), skin layer and a porous substructure support layer
-Separation of macromolecules
-Only surface deposition: no internal pore plugging; so, relatively easy to remove, irreversible

Nanofiltration (NF)

- NF Removes molecules in the 0.001 micron range
DP 0.5 to 6 MPa
MWCO: 0.2 to 200
-NF is essentially a lower-pressure version of reverse osmosis
-NF performance characteristics between reverse osmosis and ultrafiltration

Reverse Osmosis (RO)
-Membrane: similar to UF, thin active layer; porous support layer

-Operating Pressure: 1.0 - 10 MPa

-RO has the separation range of 0.0001 to 0.001mm

Electrical driven membrane processes
-Electrodialysis (ED)

Concentration driven membrane processes
-Dialysis
-Osmosis

---------------------------------------------------------
Based on Modules, membrane module refers to the device which houses the membrane element:

• Tubular membrane module

- Membrane is cast inside the support tube
- Tubular membranes have a diameter of 5 - 15 mm
- High SS tolerance
- Flow is usually inside out
-Mainly MF and UF
- Low packing density, high prices per module

• Hollow fibre membrane module

- Consists of a bundle of hundreds and thousands of hallow fiber
- Entire assembly is inserted into a pressure vessel
- Feed can be applied inside of the fiber (inside-out flow) outside (outside-in flow)
- Highest packing density of all.
- Hollow fiber is used mainly for NF and RO

•Spiral wound membrane module

- Flexible permeate spacer is provided between two flat sheet membranes
- Membrane: sealed three side and open side is attached to perforated pipe
- Flow is in a spiral pattern.
- Membrane envelop is spirally wound along with a feed spacer
- Filtrate is collected within the envelop and piped out
- Packing density:high
- RO and NF

• Plate and frame

--------------------------------------------------
Membrane Fouling: Deposition or accumulation of solids on the membrane.

Fouling causes resistance to flow through the membrane and eventual decline in overall flux.
Three major mechanisms of resistance flow:
-Pore narrowing
-Pore plugging
-Gel/cake formation due to concentration polarization

Separation processes

Separation processes based on the molecular properties are summarixed as follows:

Microbial pretreatment of piomass

Biological pretreatment of wood chips has attract great attention and research activities to reduce the mechanical pulping refining energy. The same principle and idea can be applied to biomass pretreatment for lignocellulosic ethanol production.

A review of the literature suggests that fungal pretreatment could potentially lower the severity requirements of acid, temperature and time. These reductions in severity are also expected to result in less biomass degradation and consequently lower inhibitor concentrations compared to conventional thermochemical pretreatment. Furthermore, potential advantages of fungal pretreatment of agricultural residues, such as corn stover, are suggested by its effectiveness
in improving the cellulose digestibility of many types of forage fiber and agricultural wastes.

The bottleneck for cellulosic bioethanol commercilization

When talking about the bottleneck of cellulosic bioethanol commercialization, the 1st pop-up issues may be the technology of pretreatment, the efficiency of enzymatic hydrolysis, C5 microorganisms, etc. Another factor may be too much politics in this area. Too much talk, less action; too many talkers, not many real workers.....

Hydrothermal treatment of biomass using phosphorous acid as an additive

Instead of using as a solvent, phosphorous acid can be also used as a catalyst for biomass pretreatment.

Based on the study in Japan, the yield of monosaccharide obtained from the rice straw using hydrothermal treatment was low. but when 50 mM phosphoric acid solution was used to hydrothermal system, most of xylan in rice straw was successfully hydrolyzed to xylose and some amount of glucan was also converted into glucose at 160 °C for 15 min. The maximum xylose and glucose yields were 54.1 and 15.0 % at this condition, respectively.

The remained cellulose residue after hydrothermal treatment with phosphoric acid at 160 °C for 15 min was easily saccharified to glucose by enzymatic hydrolysis. After the enzymatic saccharification, the glucose yield was 81.6 %.

After neutralization of hydrolysate with NaOH, the salt formed is sodium phosphate. This salt used as nutrient by microorganisms.

Phosphoric acid pretreatment of biomass

It has been noticed that concentrated phosphoric acid is an ideal cellulose solvent. The advantages of phosphoric acid pretreatment of biomass includes:

• cellulose dissolution by phosphoric acid occurs at low temperatures
• phosphoric acid can dissolve cellulose in the presence of water
• the regenerated cellulose remains in an amorphous form suitable for hydrolysis
• the residual phosphorous acid has no inhibitory effects on the sequential
  hydrolysis and fermentation.

But what about the cost?

Complete Fermentation of Xylose and Methylglucuronoxylose

Complete Fermentation of Xylose and Methylglucuronoxylose Derived from Methylglucuronoxylan by Enterobacter asburiae Strain JDR-1

Changhao Bi, John D. Rice, and James F. Preston*
Department of Microbiology and Cell Science, University of Florida, Gainesville, Florida 32611

Abstract:

Acid pretreatment is commonly used to release pentoses from the hemicellulose fraction of cellulosic biomass for bioconversion. The predominant pentose in the hemicellulose fraction of hardwoods and crop residues is xylose in the polysaccharide methylglucuronoxylan, in which as many as one in six of the β-1,4-linked xylopyranose residues is substituted with -1,2-linked 4-O-methylglucuronopyranose. Resistance of the -1,2-methylglucuronosyl linkages to acid hydrolysis results in release of the aldobiuronate 4-O-methylglucuronoxylose, which is not fermented by bacterial biocatalysts currently used for bioconversion of hemicellulose. Enterobacter asburiae strain JDR-1, isolated from colonized hardwood (sweetgum), efficiently ferments both methylglucuronoxylose and xylose, producing predominantly ethanol and acetate. 13C-nuclear magnetic resonance studies defined the Embden-Meyerhof pathway for metabolism of glucose and the pentose phosphate pathway for xylose metabolism. Rates of substrate utilization, product formation, and molar growth yields indicated methylglucuronoxylose is transported into the cell and hydrolyzed to release methanol, xylose, and hexauronate. Enterobacter asburiae strain JDR-1 is the first microorganism described that ferments methylglucuronoxylose generated along with xylose during the acid-mediated saccharification of hemicellulose. Genetic definition of the methylglucuronoxylose utilization pathway may allow metabolic engineering of established gram-negative bacterial biocatalysts for complete bioconversion of acid hydrolysates of methylglucuronoxylan. Alternatively, Enterobacter asburiae strain JDR-1 may be engineered for the efficient conversion of acid hydrolysates of hemicellulose to biofuels and chemical feedstocks.

Continuous cultivation process

Continuous cultivation process has been developed to answer the need of rapid ethanol production for fuels and chemical in the near future.

Batch cultivation is rarely applied for dilute-acid hydrolyzate cultivation since the cells is exposed directly to the high concentration level of inhibitors. This problem is commonly overcome by applying fed-batch cultivation where the concentrations of inhibitors in the media are controlled by regulation of the feeding. The continuous mode of cultivation has generally been of interest to minimize the total time of fermentation and increase the volumetric production rate of ethanol as well as decrease the investment cost. Because the bioreactor is fed continuously, it would be possible to avoid sudden inhibition to the yeast. Therefore, continuous cultivation mode is a suitable alternative for large-scale ethanol production.

Cell immobilization

Cell immobilization is defined as “the physical confinement or localization of intact cells to a certain region of space with preservation of some desired catalytic activity”.

The advantages of cell immobilization include-

• enhancement of fermentation productivity;
• feasibility of continuous processing;
• cell stability;
• lower costs of recovery and recycling in downstream processing.
  

The techniques can be divided into four major categories based on the physical mechanisms:

• attachment or adsorption on solid carrier;
• entrapment within a porous matrix;
• self-aggregation by flocculation (natural) or with cross-linking agent (artificially induced);
• and cell containment behind a barrier.
  

Cell entrapment is a simple and useful method to provide a simulation of high-density cells used in small-scale bioreactors where a cell recirculation method with e.g. filter and continuous centrifuge is rather difficult. But there is a need for using the cell entrapment method in industry due to additional cost of immobilization material.

Self-aggregation of cells leading to formation of a flock and easy sinking is very useful for cell recirculation, and thus performs cell retaining. Yeast has a natural flocculation capability which is usually employed in the brewing industry. The flocculation of yeast is a reversible, asexual and calcium-dependent process. Lectin-like proteins, so-called flocculins, which stick out of the cell walls of flocculation cells, selectively bind mannose residues present in the cell walls of adjacent yeast cells. Calcium ions in the medium are needed in order to activate the flocculins. Once the yeast forms flocks, the separation of cells from broth can easily be carried out by sedimentation; thus cell recirculation is performed without any force-driven separation, i.e. filtration and centrifugation.

By-products of dilute-acid hydrolysis of lignocellulosic biomass

Dilute-acid hydrolysis is still a cheap and fast process to obtain sugar from lignocellulosic materials. However, a significant drawback of dilute-acid hydrolysis is the generation of several by-products during the process.

Organic acids

A large number of aliphatic acids are present in dilute-acid hydrolyzates originated from wood extractives, lignin degradation and sugar degradation.

• Acetic acid: a major acid constituent in hydrolyzates and is mainly produced from degradation of the acetyl group in the polysaccharides
• Levulinic acid and formic acid are products of sugar degradation
• Several kinds of fatty acids such as hexadecanoic, 9,12-octadecadienoic, oleic and octadecanoic that most likely are unmodified wood extractives, in addition to short-chain and branched aliphatic acids such as 2-methyl-2-hydroxybutanoic acid, methyl propanedionic acid and methyl botanedioic acid. These last acids are not important in terms of concentration, and thus result insignificant effects to the yeast.

The undissociated acids are harmful to the cells and inhibit cell growth. They are liposoluble and thus can diffuse across the plasma membrane into cytosol and may dissociate intracellularly. In order to maintain intracellular pH, protons must be transported across the membrane by the action of plasma membrane ATPase. This results in an increase of ATP consumption, and thereby causes lower biomass yield. In anaerobic conditions, ATP generation is achieved by the ethanol production pathway, resulting in higher ethanol yield at the expense of biomass formation. However, above critical extracellular concentration of undissociated acid, the diffusion rate of undissociated acid can exceed the transport capacity of the plasma membrane ATPase, and intracellular acidification occurs. It is found that the limit of extracellular pH at different acetic acid concentrations which allow yeast to grow. It was found that growth was possible at a pH not less than of 4.7 in cultivation containing 10 g/L acetic acid. Therefore, acetic acid is innocuous if it exists in low concentration or the cultivation is carried out at a pH higher than the extracellular pH limit.

Phenolic compounds

There are a number of phenolic compounds recognized in lignocellulosic hydrolyzates, including

• 3-methoxy-4-hydroxybenzaldehyde
• 4-hydroxyacetophenone
• vanillin
• syringaldehyde
• acetovanilone
• ferulic acid
• vanillic acid
• 4- hydroxybenzoic acid

These compounds are mainly liberated from lignin degradation in addition to aromatic wood extractives. The phenol aldehydes and phenol ketones were found as the worst inhibitors. Moreover, it was also shown the low molecular weight phenolic compounds are more toxic.

Phenolic compounds are considered to be important inhibitors due to their inhibitory effect in fermentation of lignocellulosic hydrolyzates. These compounds partition biological membranes and cause loss of integrity, hence disturb their ability to serve as selective barriers and enzyme matrices. The inhibition mechanism of phenolic compounds has not been elucidated yet.

Studies in inhibitory action of phenolic compounds have been carried out using higher concentrations than are actually present in the hydrolyzates. The water solubility of phenolic compounds is limited and depends on the composition of the liquid, which is different in hydrolyzates and the defined medium; therefore it is possible that the concentration at which the microorganism suffered has been lower. In addition, S. cerevisiae assimilates vanillin, hydroxybenzaldehyde, and syringaldehyde during fermentation, while growth has been reported on cathecol, recorcinol.

Furan compounds

Furfural and 5-hydroxymethyl furfural (HMF) have been found as further hydrolysis products of pentoses and hexoses respectively. Pentoses form furfural in high yield; but if the furfural is not removed as formed, it partially condenses into high-molecular-weight materials. By an analogous process, hexoses yield HMF which, on continued heating, yields levulinic acid and formic acid. Furfural has been reported to be a strong inhibitor for S. cerevisiae. The furfural concentration above 1 g/L was found to decrease significantly the CO2 evolution rate, the cell multiplication and the total viable cell number in the early phase of fermentation. During anaerobic fermentation, reduction of furfural to furfuryl alcohol occurs with high yields, while furoic acid is produced from oxidation of furfural during aerobic cultivation. In both cases, NADH-dependent alcohol dehydrogenase (ADH) is believed to be responsible for furfural conversion in yeasts.

HMF is chemically related to furfural and thus has similar inhibitory effects as furfural, except that it has a lower conversion rate which might be due to lower membrane permeability. It is also discovered that an addition of 4 g/L of HMF decreased the CO2 evolution rate (32%), ethanol production rate (40%), and specific growth rate (70%). However, these inhibitory effects were less than those caused by the same amount of furfural, and thus HMF cannot be considered as acutely toxic as furfural for growth and fermentation of S. cerevisiae.

The conversion rate of furfural is much faster than the conversion rate of HMF. Furthermore, HMF is converted to 5-hydroxymethyl furfuryl alcohol with a similar mechanism as it was shown in the case of furfural conversion.

Another potential barrier to biofuel commercialization

Based on Brent Erickson, the executive vice president of BIO’s Industrial & Environmental Section, a cellulosic biorefinery that produces both advanced biofuels and biobased products could create as many as 2,200 new jobs and increase economic activity by more than $1 billion.

This is a good news!

However, this will not become true until a real biorefinery plant nearly take off. Before that, a series of barriers need to be overcome. In addtion to the technical, engineering, financial, and market issue, a key factor to be easily overlooked is the technical resources. We are not lack of people but we are lack of technical professionals who have been well trained with solid and sound fundemental understanding of the technology.

When biofuels become a hot spot, it seems that thousands of biofuel specialists are available in the market overnight.

However, it is not difficult to find that quite a lot of them are not real experts. Unfortunately, they are in the position to provide advices, suggestions to the government, company managers or make decision by themselves. As a result, some key technology development or technology commercilization is delayed; some companies go to wrong direction; some companies have to be closed due to the bad decision..... Just like a diagnosis for a patient from a "fake" doctor, the patient treatment was delayed, or wrong description was given to the patient. What a result can we expect from this?

As a result, the time has been wasted; the money has been wasted, the opportunities has passed; and a lot of related people got hurt, i.e. human resources was also wasted.

It is so-called 'a busines can both succeed and fail because of human resources!'

Lignin solvents may work better for biomass treatment

Lignin is the big barrier for pretreatment and enzyme hydrolysis of lignocellulosic biomass. The idea to use ionic liquids is to decrystallize of the cellulose to make it more digestible for enzymes.

If we can use lignin solvent like dioxine to break down lignin linkages, it may significantly increase the enyzme digestability and reduce enzyme loadings during enzyme hydrolysis.

The misundstanding of lime treatment for detoxification of lignocellulosics hydrolyzate

Traditionally, lime treatment was used to remove phenolic compounds in pulping acid sulfite pulping spent liquor because of the formation precipitate calcium lignosulfonate.Unfortunately, this method was also proposed today to be used widely to remove other by-products such as acetic acid, furfural, and HMF in the hydrolyzate, which is not right from fundamental aspect. These by-products can be removed due to the lime trap from lime treatment, which also traps sugars and leads to 20-35% sugar loss.

Two science research progress on biofuel plants

These two progress means more than the research itself. It will significantly impact the lignocellulosic biomass biofuel production process, equipment, and cost.

1. Modified Plants May Yield More Biofuel.

2. Modified Lignin Has Potential Benefits For Ethanol, Paper And Feed.

The challenge of IL pretreatment of biomass

The challenge of IL pretreatment of biomass:

1. Moisture of biomass. Some level of water will reduce the efficiency and rate of decrystallization of cellulose, what is the tolerance of water level of biomass for IL function?

2. Particle size reduction. Lignocellulosics is the complex of lignin, cellulose, and hemicelluloses and is more difficult to degrade than cellulose due to strong interactions between lignin and glucose. So, particle size reduction is needed before IL pretreatment

3. IL recovery. After treatment, water is usually added to extract IL out. The question is whether we can extract 100% of the IL used out of the sugar solution by simple extraction. The IL lost in the sugar will cause 1) the cost increase of IL usage 2) the potential toxicity on microorganism during fermentation because IL is salt

4. Separation of sugar and IL. Since some ILs is miscible with water, sugar will dissolve in ILs. The separation of sugar will become a cost and technological issue.

5. The issue of hemicelluloses and lignin. How to handle these two compounds?

Efficiency and cost consideration for detoxification of lignocellulosics hydrolyzate

The hydrolyzate of lignocellulocis need to be detoxified before fermentation. However, there must be a consideration of the efficiency and cost when a method is selected.

Biological and membrane are very effective but not cost-effective for bioethanol production;
Lime treatment is effective and cheap but the sugar loss and by-product (gypsum) handling is another cost and environmental issue.

I like the idea of stripping combined with activated charcoal treatment, which works well with good cost advantage. The charcoal can be easily re-activated and reuse.

A list of Biodiesel Kits and Production Equipment suppliers

Websites:

1. http://journeytoforever.org/biofuel_supply.html
2. http://www.biodieselnow.com/blogs/methods/archive/2007/01/27/the-basics-of-biodiesel-production-101.aspx
3. http://www.biodieselpictures.com/

Suppliers:

1. Alvan Blanch is a British manufacturing and project engineering company with a global outlook, specializing in the design, production and supply of quality machines and integrated systems for the primary and secondary processing of agricultural produce and waste materials.

2. Arbor Biofuels Company LLC, is an Ann Arbor based start-up company. We manufacture methyl ester (biodiesel) from the collected vegetable oil waste of our area. We also specialize in the fabrication Biodiesel Purification equipment, particularly in the area of ion exchange columns or dry wash towers.

3. Anyang General International Co., Ltd (AGICO) is specially in design and manufacture of complete oil plants and equipments, importing and exporting various mechanical products and engaging in external economic and technical cooperation.

4. ARBODS Ltd supplies fuel/energy saver means/equipments and a complete range of biodiesel processing equipments, reactors, plants. From simple compact processing kits to full-scale commercial plants with equipment, personnel and training provided as required.

5. BDC Systems is a private company with many years experience in grain drying, cleaning, storage and handling equipment, also milling and mixing.

6. Bio100Supply ,located in Georgia, U.S.A. provides all of the lab ware, pumps, safety equipment and chemicals that are necessary for making homemade biodiesel or creating a SVO conversion kit for your car.

7. BioMac, supply a complete range of biodiesel processing equipment at an affordable cost. BMC provides a complete range of biodiesel processors and support system from compact units to full-scale industrial plants, tailored to customers’ needs.

8. Bio Development, LLC provides quality products to consumers searching for a way to produce alternative fuels in order to create a cleaner, safer environment.

9. Biodiesel 123. Biodiesel123 is made up of a team intent on manufacturing the safe and efficient biodiesel processors available.

10. Biodiesel Experts International, The premier source for Biodiesel process equipment, engineering, sales and support.

11. Bio-Diesel International, a company in Austria provides solutions for the industrial use of renewable resources.

12. Biodiesel Logic Inc is based on the vision that small to medium scale biodiesel production is not only logical but profitable. Also that small to medium scale packaged processing equipment can be improved by applying industrial engineering criteria that has not been normally available to the small biodiesel producer.

13. Orbitek Biodiesel Production Equipment specializes in Biodiesel Producers, Biodiesel Productions, Biodiesel Production Systems, Biodiesel Technology and Renewable Fuels.

14. Biodiesel Solutions manufactures the BiodieselMaster® line of continuous-flow automated processors at its 30,000 sq. ft. factory in Sparks, NV. It is an engineering company dedicated to supporting small-scale biodieselers in every aspect of making their own fuel. This company also supplies materials to biodiesel cottage industries and biodiesel plants.

15. Biodiesel-technologies's proprietary process offers advantages and cost efficiencies over its competitors for technology, production, and facilities in this emerging industry.

16. Biodiesel Technologies International, LTD (Biodiesel Gear) -Located in Illinois, this company produces cost effective biodiesel p equipment and and all of the accessories that you wil lneed to create your own homemade brew. They also supply individual parts such as hoses and pumps to home brewers who are attempting to design a one of a kind biodiesel processor.

17. Biodiesel Warehouse is focused on providing a practical online resource for the small to medium capacity (5 to 1 million gallons per year) home brewer, coop, farm or business. We believe in using high quality materials, and only sell processors which utilize steel plumbing and steel reaction vessels.

18. Biofuels, an Argentina company that provides pressurized steel units designed for individual or coop use. They will process both virgin or waste vegetable oil, and are fully self-contained.

19. Cavitation Technology Inc., is an engineering and manufacturing company, specializing in the production of the biodiesel equipment with its signature product.

20. Crown Iron Works Company's biodiesel production equipment is designed for continuous operation for maximum efficiency and safety. The process is a two stage transesterification reaction followed by ester washing, drying and alcohol recovery. Excess methanol is recovered from all product streams to virtually eliminate discharge to air and water and allow its reuse in the process.

21. DirectIndustry is the permanent virtual exhibition, connecting manufacturers and buyers in 5 languages, around the world. With 4 million visitors a month sourcing from over 48,000 industrial products, it is the biggest industrial exhibition online.

22. Ascension Industries, Inc. - Durco Industrial Filtration Division provides their state-of-the-art Biodiesel Dry Washing & Filtration Systems in the US,Europe, and Asia.

23. Doctor Diesel, Located in California, this company specializes in producing small-scale biodiesel making equipment and parts for the hobbyist or serious home brewer. It is also a good site to get advice on making your own biodiesel.

24. UkrBudMash Ltd., a company in Ukraine to provide BioDieselMatchTM continous stream biodiesel production systems.

25. Energea, a company in Austria with CTER technology "Continuous Trans Esterification Reactor" that opens a new chapter in biodiesel production: With up to 50% lower costs of investment and practically 100% yield. Turn-key installations, built in container sized modules, provide for considerably less space requirement.

26. EuroFuelTech manufactures all of the biodiesel equipment needed to produce quality fuel, whether you want to make 2,000 litres per day or 400,000 litres per day.

27. Evolution Biodiesel,LLC provides biodiesel processors.

28. EZBiodiesel is a company providing a full range of solutions to your biodiesel processor needs. It is ready to run processors from 20-800+ gallons, & Do It Yourself Kits available as well as parts & accessories and also has a vast amount of Free information about biodiesel.

29. Fertile Fuels is a Colorado based research and fuel production company focused on rural and urban Biodiesel production.

30. GHP-Biodiesel, a company in Germany provides small scale biodiesel equipment plus logistic support. Renting of equipment.

31. Green Fuels provides small and medium scale biodiesel equipment in Europe.

32. Green Grid Solutions, a Canadian company provides Biodiesel manufacturing supplies.

33. Greenline Industries is a proven provider of innovative technologies for the production of biodiesel fuel. It manufactures award-winning biodiesel production equipment and provide engineering and consulting services for the construction and management of biodiesel facilities.

34. GreenShift develops and commercializes clean technologies that facilitate the efficient use of natural resources.

35. Goat Industries, a company in UK to provide biodiesel equipment, 'veggie boost'er, conversion kits etc.

36. Home BioDiesel offers a quality product for the conversion of vegetable oils into diesel fuel.

37. Homebiodieselkits provides complete biodiesel kits assembled and ready to produce biodiesel fuel the same day. Including a superior fuel filtering system and solid one piece steel frame.

38. IBG Monforts Oekotec can certainly rightfully claim to be an expert in the oil seed processing industry.

39. Imerjent LLC, a start-up technology firm, launched in January 2007 to bring an innovative, highly automated biodiesel production system to market. The firm will specialize in manufacturing cost-effective, medium-scale modular systems.

40. JatroDiesel is a Biodiesel Production Equipment provider and Biodiesel producer.

41. KEE CHEUNG CO, located in China, is biology diesel-oil equipment factory, with strong technology strength, involved in biodiesel research, product, biodiesel equipment manufacture.

42. Kemia Gmbh Manufactures Biodiesel Production Equipment And Waste Oil Recycling Equipment. Containerized Units, Quick Delivery, Quick Installation.

43. Lurgi provides proprietary technologies and exclusively licensed technologies in the areas gas-to-petrochemical products via synthetic gas or methanol and synthetic fuels, gas generation and treatment, petrochemical intermediate and end products, biofuels as well as food and oleochemicals.

44. National Tank Outlet is your source for plastic storage tanks.

45. Nova Biosource Fuels, Inc. is a Leading Edge Provider of Biodiesel using its Proprietary Patented Process Technology. Nova currently has over 230 million gallons of Biodiesel in various stages of construction or in operation.

46. NAZCO distributes renewable alternative fuels; bulk vegetable oils and biodiesel production equipment for a cleaner world.

47. Olympia Green Fuels LLC, located in Washington, this company sells a modular kit for making biodiesel at home. These kits produce biodiesel from both fresh and waste vegetable oil. These processors will convert both fresh and waste vegetable oil into a washed and polished biodiesel fuel that is capable of meeting the quality standards of the ASTM.

48. Pacific Biodiesel, Inc. manages biodiesel plants in Hawaii, Oregon and Texas and creates truly sustainable, community-based biodiesel production facilities that incorporate feedstock usage, fuel processing and product distribution within localized geographic areas in order to maximize the economic and environmental benefits and minimize energy consumption.

49. PESCO-BEAM Bio-diesel Systems are the most complete systems on the market today.

50. Planet Biodiesel, located in Colorado this company supplies the information and products you need to make your own biodiesel fuel.

51. Renewable Energy Group (REG) leads the nation in biodiesel production and marketing.

52. SE-Energy specializes in offering a single-source solution for 40mm-320mm gal/yr biodiesel plants.

53. Special Technologies, located in Ukraine and provides modern technologies and equipment for biodiesel production.

54. SunBio Systems (SB) is focused on the small to mid sized biodiesel equipment market. SBS biodiesel machines range from the university grade research and development system to truck fleet and rural area systems, to mid sized plants that produce 50 million gallons per year.

55. Utah Biodiesel Supply carries a large selection of Biodiesel home brewing supplies, equipment, literature, soap, bumper stickers, decals, and more.

56. Wintek Corporation has methanol recovery systems for Biodiesel and glycerin to meet ASTM standards. Custom design in carbon or stainless steel for capacities of 5-150GPM.

57. YiXing HuaDing Food Machinery Co,.Ltd, a company in China and is able to provide design, manufacturing, fixing, debugging of 10 t/d-600t/d oil refining production line and main equipments, including disc centrifuge, bleaching tower, deodorization tower.

New Opportunities from ionic liquids in biorefinery

3rd Generation Biomass Conversion Technologies can be developed by using ionic liquid pretreatment of the lignocellulosic biomass. The recalcitrance of biomass due to crystallinity and lignin sheathing of biomass will be overcome leading to enhanced yields. The enzymatic hydrolysis of cellulose pretreated with ionic liquids will be faster and more efficient.