Newsfront: Teaching an Old Dog New Tricks

April 15, 2008 | By Joy LePree

While it may be one of the oldest chemical processes known to mankind, fermentation is getting a high-tech face lift as industrial biotech firms put a fresh spin on the age-old technology in an effort to develop advanced products in a more environmentally friendly and less costly way.

"The technology is generally the same — using microbial systems and metabolic fermentation — but by applying genetic engineering we are able to improve the microbes used in that fermentation. We are also using enzymes produced by these genetically enhanced microbes to ferment starch more effectively," explains Brent Erickson, executive vice president of the industrial and environmental section at the Biotechnology Industry Organization (BIO; Washington, D.C.).

The bioplastics, biofuels and biopharmaceuticals industries are increasingly taking advantage of fermentation because it helps control production costs and is easier on the environment, according to the experts.

"The area of bioprocessing is now extremely important in terms of finding ways to control production costs and finding new methods to recover molecules," says Mani Subramanian, director of the Center for Biocatalysis and Bioprocessing at the University of Iowa (Coralville). "Producing the molecules isn’t the difficult part," he says. "The problem comes when we try to recover them from the media. And, as the science behind industrial biotechnology advances and we find more effective ways to do this, it will begin to help control costs."

As for being environmentally friendly, BIO claims that bioprocessing tends to prevent pollution in the first place, reducing — and possibly eliminating — the need for pollution control in certain processes.

"Industrial biotechnology is already reducing pollution and manufacturing costs in some industry sectors," says Erickson. "It provides a new set of tools that hold great promise to further reduce pollution and the consumption of raw materials if deployed more broadly. This, in turn, can reduce the cost of producing goods and may lead to better products."

For example, BIO says that biotechnology process changes in plastic production replace petrochemical feedstocks with ones made from organic materials, such as corn, which could reduce the demand for petrochemicals by 20 to 80%. Because bioplastics are biodegradable, their use could also reduce plastics in the waste stream by up to 80%. Waste burdens are reduced partly because disposable food service items such as plates, cups and containers can be composted along with food waste, eliminating the need for separation. These plastics can also be used to make many other products, ranging from clothing to car parts, which can be composted instead of disposed of in landfills.

BIO says that more than 80-billion lb of plastic products are produced annually in the U.S. Of that, 1-billion lb are bio-based. The remaining potential for environmental benefits and reduced demand for foreign oil is obviously substantial. For example, if all plastics were made from bio-based polylactic acid, oil consumption would decrease by 90 – 145-million bbl/yr, which is about as much oil as the U.S. consumes in one week.

The results in biopharmaceuticals are equally impressive. For example, biotechnology process changes in the production of riboflavin reduce associated carbon dioxide emissions by 80% and water emissions by 67%, says BIO. Changes in the production of the antibiotic cephalexin reduce carbon dioxide emissions by 50%, energy demand by 20% and water usage by 75%.

In biofuels, biotechnology process changes allow for bioethanol production not only from corn, but also from cellulosic biomass. BIO says the energy demand during production of ethanol in this way falls by almost 3%. According to the organization, the closed-loop nature of using cellulosic biomass to produce bioethanol can contribute substantially to the mitigation of greenhouse gas emissions and provide a partial solution to global warming (however, see pp. 21 – 22). The U.S. has the potential to produce between 20 and 40-billion gal of bioethanol from cellulosic biomass in the future.

Moving into new territory

Because of the major R&D and equipment investments necessary to bring bio-based products to market, it’s the big chemical firms, such as Cargill, Archer Daniels Midland, DSM and DuPont, that are throwing capital at the technology, especially when it comes to bio-based plastics.

For example, the largest commercial producer of bioplastics in the U.S. is NatureWorks LLC (Minnetonka, Minn.), which is a joint venture between Cargill (Minneapolis, Minn.) and Teijin Ltd. (Tokyo, Japan). At the NatureWorks plant in Blair, Nebraska, the company uses corn sugar to produce PLA plastic packaging materials, as well as fibers. PLA plastic, which offers characteristics similar to petrochemical-based plastics and can be processed on the same equipment as conventional plastics, is made by fermenting starch from crops, such as corn starch, into lactic acid that is then polymerized.

Archer Daniels Midland Co. (ADM; Decatur, Ill.), which specializes in agricultural processing and fermentation, has teamed with Metabolix Inc. (Cambridge, Mass.) to produce a bioplastic called Mirel. The bioplastic commercial manufacturing facility is located adjacent to ADM’s corn wet mill in Clinton, Iowa. Produced from renewable resources like corn sugar, Mirel provides an alternative to traditional, oil-based plastics for use in products as diverse as cosmetics, food industry packaging and consumer goods, says Oliver Peoples, co-founder and chief science officer at Metabolix.

Peoples says his firm has developed an advanced production technology which makes bioplastics more effectively and efficiently in a process that is scaleable to large scale. "What’s different from conventional plastics is that these are made of sugar, which is a renewable resource," explains Peoples. "Another important aspect is that they provide a completely bio-based solution to a material that is very much a generator of greenhouse gases." Peoples says Mirel has the potential to reduce petrochemical usage by about 95% and greenhouse gas production by 200% while providing the same functional performance as petroleum-based plastics.

"The only difference between bio-based plastics and conventional plastics is that these are bioproduced in a cleaner way and are completely biodegradable at the end of their lifecycle," says Peoples.

In another collaborative move, DuPont (Wilmington, Del.) has teamed with Genencor International (Rochester, N.Y.) to bioproduce 1,3-propanediol, a key monomer used for the production of DuPont Sorona polymer and fiber. The polymers have environmental benefits since the manufacturing process begins with glucose from corn and the finished Sorona is recyclable.

DuPont also has a hand in the biofuels till, as well. The chemical giant is working with the U.S. Dept. of Energy (DOE; Washington, D.C.) to fund a four-year research program to develop technology to convert corn stover into ethanol. In addition, DuPont is involved in a biobutanol partnership with BP and Advanced Biofuels Pipeline. The partnership with BP (Warrenville, Ill.) to develop biobutanol is intended to bring advanced biofuels to market to expand the use of biofuels in gasoline. Biobutanol will be the first product available via the partnership and is said to enhance ethanol-gasoline blends by lowering the vapor pressure when co-blended with these fuels. It also enhances fuel stability of biobutanol-gasoline blends, giving it the potential to be distributed via the existing fuel supply infrastructure.

Technological advances

Despite the financial backing of the chemical giants, bioprocessing advancements would not be possible without fermentation equipment. But because much of the work in bioprocessing is in the early developmental stages, researchers are still trying to figure out exactly which techniques and types of equipment work best.

For example, while experts still don’t know with certainty which biofuel will be the one that is ultimately chosen for common use, they do know that the process will need to be a clean one. For this reason equipment that is engineered to be sterile is currently getting attention, says Sue Reeb, product manager and staff scientist at GEA Niro (Columbia, Md.) "There’s currently a lot of activity in this sector trying to determine THE technology," says Reeb. "There’s a lot of work at the lab scale with fermenters and bioreactors and the cellulosic materials. But these processes don’t necessarily lend themselves to being sterile or clean."

Reeb says that in the beginning researchers weren’t really worried about biofuel cleanliness, but that’s changing because they’ve learned that contamination affects yields.

"Since we engineer cleanliness into the process and do a lot of hygienic design as a consequence of our dairy experience, we are getting calls from companies that are building pilot plants for these processes," says Reeb. She adds that sterilization was used on a small scale, but it’s not energy efficient to do a lot of steam sterilization on a larger scale. "They are turning to us to build a hygienic design into the bioreactors, fermenters and, ultimately, the plants," she says. "This ensures that there’s cleanliness by design."

According to Christian Stoffers, marketing manager of natural resources with Alfa Laval (Richmond, Va.), there are additional ways to improve yield in biofuels production. "Today most bioethanol plants work on the same principle as normal potable alcohol plants. It is really finding the right yeast cells that provide the most benefits regarding yield improvements," Stoffers says. "However, certain adjacent technologies in the equipment can make a significant difference in the future."

Efficient cleaning of fermenter vessels provides a good environment for the yeast and avoids infections, which will lower the activity of the yeast and will decrease yield. High-speed separators and membranes can also be used to purify the product and get rid of substances that can lower the activity of the yeast. And, because the fermentation process achieves the best yield within a certain temperature range, a well-working heat exchanger can ensure that the fermenter is kept within the proper range.

Alfa Laval’s fermenter cooler, Widegap350, provides a solution to these needs. The unit can handle fibrous materials found in grain-based processes and achieves a high thermal efficiency with regard to a tight temperature approach between the warm mash and the cooling water.

While the biofuel industry is busy figuring these things out, biopharm is developing a different set of requirements. "Processing bioactive materials is about more than moving volume through a system. It’s about meeting production goals with products that are active, intact and undamaged," notes Ian Sellick, director of marketing at Pall Life Sciences (East Hills, N.Y.). "But traditional tangential flow filtration and centrifuges don’t always accomplish this, particularly when processing viscous or high-solids materials."

As a result, Pall launched the PallSep Biotech vibrating membrane filter system, which is designed for processing very difficult materials. The system offers better flux and capacity compared to static filtration. It employs vibrational energy to generate shear force on the order of 1,000 to 150,000 s ^–1 at the membrane surface, which reduces the effects of membrane fouling and permits gentle processing and high recovery rates. The system is effective in high-solids and high-viscosity applications where heat can’t be applied, such as the recovery of therapeutic proteins.

Another issue common to biopharmaceuticals is the lack of willingness to invest large sums of capital during the early developmental stages. "For this reason, we are working to develop disposable technologies for all the unit operations in processes," says Thomas Scholz, marketing director with Pall Life Sciences. "For example, our Kleenpak TFF Micorfiltration Capsule offers a high-flux rate with minimal hold up volumes, even under demanding bioprocessing conditions. Biopharmaceutical manufacturers benefit from the unique combination of an efficient microfiltration mechanism packaged in a disposable, easy-to-use capsule."

While each segment has special needs, there is one common platform from which all industrial biotech processes can benefit. "The optimization and automation of processes through software integration of multiple analyzers, probes and ancillary equipment is now possible," says Richard Mirro, product manager for autoclavable fermenters and bioreactors with New Brunswick Scientific (Edison, N.J.). "Fermentation may be one of the oldest sciences in the world, but now we are better able to control it and gather more data than we could years ago thanks to the addition of software that allows users to link analyzers and testers into one data system. This helps end users examine their processes and data more carefully in real time," says Mirro. "This is where the real advances have been made."

Subramanian agrees. "Software made for monitoring the fermentation and process analyticals has made significant leaps. Whether for biotherapeutics or biofuels, you can monitor how the organisms are growing, what gases are coming out of the equipment and change the fermentation process accordingly. Advanced software has helped maximize all the processes," he says.

And, this is helping to put bioprocessing on the map as a feasible, practical, cost-effective and environmentally friendly chemical process.

CONSUMER PRODUCTS MADE WITH BIOTECHNOLOGY (SOURCE: BIO)
Product	Old manufacturing process	New industrial biotech process	Biotech enabling technology	Benefit
Detergent	Phosphates added as a brightening and cleaning agent	Addition of biotech enzymes as brightening and cleaning agents Protease enzymes remove protein stains Lipases remove grease stains Amylases remove starch stains	Genetically enhanced microbes or fungi bioengineered to make enzymes	Elimination of water pollution from phosphates Brighter, cleaner clothes with lower temperature wash water Energy savings
Bread	Potassium bromate, a suspected cancer-causing agent at certain levels, added as a preservative and a dough strengthening agent	Addition of biotech enzymes to Enhance rising Strengthen dough Prolong freshness	Genetically enhanced microorganisms to produce baking enzymes (Directed evolution and recombinant DNA)	High quality bread Longer shelf life Eliminates use of potassium bromate
Polyester bedding	Polyester produced chemically from petroleum feedstock	Biotech polyester (PLA by Cargill) produced from corn sugar feedstock Biotech polyester (PDO DuPont)	Existing bacillus microbe used to ferment corn sugar to lactic acid. Lactic acid converted to a biodegradable polymer by heating. Polymer made into plastic products and polyester	PLA polyester does not harbor body odor like other fibers Biodegradable Not made from petroleum Does not give off toxic smoke if burned
Vitamin B ₂	Toxic chemicals, such as aniline, used in a nine-step chemical synthesis process (hazardous waste generated)	One-step fermentation process uses vegetable oil as a feedstock and sugar as nutrient	Genetically enhanced microbe developed to produce vitamin B ₂ (Directed evolution)	Biologically produced without chemicals Greatly reduces hazardous waste generation and disposal
Stone-washed jeans	Open-pit mining of pumice. Fabric washed with crushed pumice stone or acid	Fabric washed with biotech enzyme (cellulases) to fade and soften jeans or khakis	Textile enzymes produced by genetically enhanced microbe (Extremophiles and recombinant DNA)	Less mining Softer fabric Reduced energy consumption Lower cost
Paper bleaching	Wood chips are boiled in a harsh chemical solution to yield pulp for paper making	Use of enzymes to selectively degrade lignin and to break down wood cell walls during pulping	Wood bleaching enzymes produced by genetically enhanced microbes (Recombinant DNA)	Reduces the use of chlorine bleach and reduces toxic dioxin in the environment Cost savings due to lower energy and chemical costs
Ethanol fuel	Food and feed grains are fermented into ethanol (a technology that is thousands of years old)	Cellulase enzyme technology allows conversion of crop residues (stems, leaves, straw and hulls) to sugars that are then converted to ethanol	Genetically enhanced organism developed to produce enzymes that convert agricultural wastes into fermentable sugars (Directed evolution, gene shuffling)	Renewable feedstock — reduces greenhouse gas emissions Increases domestic energy production Production more energy efficient than old process
Antibiotics	Chlorinated solvents and hazardous chemicals used to produce antibiotics through chemical synthesis	One step biological process using direct fermentation to produce antibiotic intermediate for cephalexin production (DSM)	Genetically enhanced organism developed to produce the key intermediate of certain antibiotics (Recombinant DNA)	65% reduction in energy consumption Overall cost savings
Contact lens solution	Surfactants or saline solutions (do not remove protein deposits) used to clean lenses	Protease enzymes remove protein deposits from the contact lens	Genetically enhanced microbes engineered to make protease enzymes (Directed evolution)	More effective contact lens cleaning Less eye irritation