science/teehnology
NEW AGE PAPER AND TEXTILES Fungi, enzymes, and closed-loop catalysis offer environmental, economic gains in manufacturing and recycling Mairin B. Brennan C&EN Washington
T
he industrial road from cotton boll to blue jeans or wood to paper wends along routes that are energyintensive or environmentally unfriendly. Expensive cleanup tolls also are inescap able—industry must bear the economic burden of continually recovering or dis posing of waste materials deemed toxic or otherwise harmful to animal life or the environment. Not surprisingly, the push is on to im prove paper and textile production. Researchers are finding ways to use bioprocessing and biotechnology to make paper and textile processing more envi ronmentally friendly and energy efficient. "We want to avoid environmental problems rather than recover from a sit uation where environmental problems have been caused," says Thomas E. Ham ilton, director of the Forest Products Lab oratory (FPL), in Madison, Wis. "The fo cus has shifted from cleaning up after ward to avoiding problems in the first place." FPL is a unit of the Department of Agriculture's Forest Service. Much of the solution lies in achieving easier access to cellulose, the major struc tural polymer in trees and plants. This long, linear glucose polymer is the stuff paper and most natural fabrics are made from. Nature rig- B\ orously shields it from harm by enclosing it in a &Ί mix of barrier polymers. fj Breaking down na ture's barricades to get -|| to the structural cellu- %\ lose fibers traditionally | entails mechanical beat ing or harsh chemical 5| treatment of cellulose source materials. How ever, hints of change are in the air. Gentler and more efficient ap
proaches to wood-pulp and paper pro cessing are being cultivated. Alongside these developments, efforts to redefine cotton and flax processing are gathering steam. In wood, cellulose fibers are protect ed in a matrix of lignin, a phenylpropanoid polymer. In cotton and flax, the fibers are protected by pectin, a methoxylated polygalacturonic acid. These two guardian polymers are fortified with a packing of hemicelluloses (heteropolysaccharides with a backbone of either xylose or glucose and mannose units). Major steps on the industrial route from tree to paper include removing the bark, chipping the wood, pulping the chips, and bleaching the pulp. Wood chips are pulped either mechanically or chemically. Mechanical pulping is energy intensive, removes very little lignin, and, as a result, yields low-quality paper. (Lig nin makes up 20 to 30% of wood, de pending on the type of tree; cellulose, about 45%.) Consequently, pulping is dominated by kraft pulping, the major chemical pulping process, which uses sodium sul fide and caustic soda. This approach re moves enough lignin to produce paper strong enough to make grocery bags. Kraft processing produces stronger pa per at a low cost in energy but "loses
50% of wood and is polluting," says mi crobiologist Masood Akhtar, vice presi dent of research and development at Biopulping International Inc., Madison, Wis. By comparison, wood loss in me chanical pulping is no more than 10%. The residual 5% lignin in chemical pulp from kraft processing is the stuff that colors the grocery bags brown, and it must be bleached out to produce white printing-quality paper. Traditional ly, the residual lignin is oxidized with chlorine or chlorine dioxide, a step that has caused environmental controversies for the paper industry. Novel bioprocessing approaches and new catalytic developments now are paving the way for a new era in wood-topaper processing. Fungi have been enlist ed to clip through lignin in wood chips to provide a softer substrate for mechan ical pulping. A self-buffering inorganic catalyst that converts residual lignin to carbon dioxide and water has been de veloped. An enzyme-based pulp-bleach ing process is in pilot plant scale-up. And an enzyme-based deinking method that can remove ink from mixed office waste, newsprint, and magazine stock has been commercialized. The use of fungi to pretreat wood chips is dubbed biopulping. Ten years in the making at FPL, the process recently has been scaled up to handle a 50-ton chip load. "Back in 1987, we formed a consor tium of FPL; the University of Wisconsin, Madison; the University of Minnesota, Minneapolis; the [nonprofit research funding agency] Energy Center of Wis consin; and 23 pulp and paper compa nies from the U.S. and abroad to screen fungi for their ability to chew up lignin in a variety of woods," Akhtar says. (Pre viously employed by the University of Wisconsin's biotechnology center, Akhtar led the biopulping effort until 1996.) In dustry, which supported the effort, was looking for ways to save on energy costs and improve pa per quality in mechanical pulping, he explains. After screening sever al hundred species, the fungus seekers got lucky. They discovered that Cenporiopsis subvermispora, a white rot fungus that's a common inhabitant of Wiscon sin's forests, chops up lignin very nicely in sever al different types of wood. MARCH 23, 1998 C&EN 39
science/technology Polyoxometalate oxidizes lignin to CO2 and H2O
Cellulose is a polymer of glucose units connected by p-l,4-glycosidic linkages OH
ta
HO O ^ CH2OH
CH2OH -Ο
HO
OH HO 0
OH
Ό CH2OH
'4 HO
CH2OH ~0 OH
[SiVgWioO^f C02 and H20
Lignin
Oxidized lignin
With this fungus in hand, FPL devel oped a simple biopulping process: Wood chips are steamed toridthem of native microorganisms, then cooled and inocu lated with a suspension of G subvermispora in a mixture of water and corn steep liquor. The liquor, a by-product of the starch industry, furnishes a growth medium for the fungus that dramatically reduces the amount and thus the cost of fungus needed to colonize the chips. "We have gone from using 3 kg to using 0.25 g to treat 1 ton of wood chips," Akhtar says. "That's well within [a desir able] economic range." The liquor itself is inexpensive—about $55 a ton—and readily available, he notes. Piles of inoculated chips are ventilat ed with filtered, humidified air, which maintains the temperature the fungus needs to be optimally active by removing the heat it generates in degrading lignin. The air also sweeps carbon dioxide away from the fungus and delivers oxygen to it. Just how C subvermispora attacks lig nin is not well understood, notes FPL re search chemist Kenneth E. Hammel. "So far as white rot fungi are concerned," he says, "lignin is just the wrapping on the candy bar. What the fungus really wants is the cellulose, which has an ordered struc ture and gives you the same piece each time you cut it. So it's good to eat. "Lignin is very amorphous," he con tinues. "Its role is to protect cellulose and hemicelluose from microbial attack. Whatever mechanism C. subvermispora uses to degrade the stuff, it has to take into account the fact that lignin is a wa ter-insoluble polymer, that it's stereoirregular, and there are no large pores in it. You can't have a situation where a typical biological enzyme that uses a careful fit to the active site is operating, because lignin doesn't have any structure like that." To explore the mode of action of C. subvermispora, Hammel synthesized various models of lignin, labeling func tional groups with carbon-13. After incu bating the polymers with fungus, he 40 MARCH 23, 1998 C&EN
used nuclear magnetic resonance [SiV2W10O40]7· spectroscopy to examine the 02 changes in the labeled groups. His results indicate that the or In a two-step, closed-loop process, the selfganism uses a 1-electron oxida buffering catalyst [Sf^W^J 6 " oxidizes lig tion mechanism to cleave the lig nin in wood pulp to carbon dioxide and wa nin substrate at select carbonter. In the anaerobicfirststep, the catalyst is carbon and carbon-aryl oxygen reduced to [SiV^W^O^7- and lignin is par bonds [Appl Environ. Microbiol, tially oxidized Addition of oxygen reoxidizes the catalyst, which completes the degra 63, 4435 (1997)]. "We don't actu dation of lignin. The pH of the reaction is ally know what the 1-electron oxi between 8 and 9, allowing cellulose to be re dant is," he says. covered undamaged from the pulp. C. subvermispora lacks lignin peroxidase, an enzyme that oper ates via a 1-electron oxidation mechanism in some other white rot fun that paper produced from biopulped gi. None of the lignin-peroxidase-contain- chips is stronger than paper derived ing fungi found so far are good candi from mechanical pulp. Akhtar and his co dates for biopulping, says Hammel, be workers have estimated that mills that produce paper for magazine printing cause they tend to degrade cellulose. During FPL's two-week biopulping could save $13 per ton of pulp and retain process, the linkages that connect lignin paper quality by shifting from the cus subunits are being cleaved, Hammel sug tomary blend of 50% mechanical pulp gests. "That's what's making the wood and 50% kraft pulp—which is blended soft." Biopulping is stopped well before with mechanical pulp to strengthen the the fungus can attack cellulose, which paper—to 55% biomechanical pulp and takes place no sooner than six weeks af 45% kraft pulp. Kraft pulp is very expen sive—$600 to $900 per ton compared ter inoculation, he says. Test runs at FPL's facility have shown with $350 to $450 per ton for mechani cal pulp. Fungal pretreatment of wood chips would cut the energy cost of mechanical pulping 30%, a savings of $10 per ton of pulp produced, says Akhtar. In a conven tional mill making 300 to 1,000 tons of wood pulp a day, that's a substantial sav ings, he notes. Alternatively, with fungal pretreat ment, the mills could increase produc tion 30% without expending additional energy. One paper company, Consolidat ed Papers, Wisconsin Rapids, Wis., has tested the biopulping process and con firmed FPL's findings, Akhtar says. The cost of incorporating the fungal approach into existing mills is minimal, he says. Constructing a facility for treating the chips would be the only expense. Biopulping also promises to deliver spin-off recycling benefits: It is being ex plored as a way to produce paper from Fungi growing on a dead tree secrete enzymes that degrade lignin and cellulose. wood waste—which would relieve some
of the burden on landfills—as well as from nonwoody plants to conserve trees. "Biopulping International and FPL are now working at biopulping kenaf, jute, straws, and other nonwoody plants," Akhtar says. Kenaf, a fast-growing annual plant grown in four states in the southern U.S., will be used for paper production. The first mill to take on this venture is scheduled to be up and running next year in Texas, he says. In collaboration with FPL, Biopulping International also is extending the biopulping approach to kraft pulping. The collaborators believe it will increase the pulp yield, reduce both the cooking time and operating temperature, and cut chemical and effluent loads as well as emissions from the kraft process. Biopulping International "is ready to market the biopulping technology," Akhtar says. Toward that end, the company is collaborating with firms that will license the technology, provide the fungal inoculum on a commercial scale, manufacture equipment for mechanical pulping, and contribute technical expertise. Another step in paper processingpulp bleaching—may soon see its traditional formula changed. Just about ready for pilot-plant testing is an environmentally friendly polyoxometalate (POM) catalyst (SiV2W10O40) that oxidizes lignin to carbon dioxide and water in a closedloop cycle. The catalyst is designed to replace a chlorine bleach step. "It's a new technology for the 21st century," says Rajai H. Atalla, head of chemistry and pulping research at FPL and a professor of chemical engineering at the University of Wisconsin. "I genu-
Akhtar: biopulping is ready to market
Lignin is derived from 4-hydroxycinnamyl alcohols CH2OH
H
3
C 0
O^OV{ H
HCJ \ ^ ^
J^
„OCH 3
CH2OH
ÇHOH
/ HoCO
ÇHOH
OCHo
CH2OH
Coniferyl alcohol (4-hydroxy-3-methoxycinnamyl alcohol)
CHoOH Source: Kenneth E. Hammel, Forest Products Laboratory
Schematic shows major linkages between phenylpropane units in softwood lignin. Most commonly, the side chain β-carbon of one unit is ether-linked to the 4-aryl position of another. Hardwood lignins are similar, but contain vary ing quantities of 3,5-dimethoxylated aromatic rings. The white rot fungus Ceriporiopsis subvermispora is thought to break the bond between the side chain a- and β-carbons as well as the bond between the side chain β-carbon and the aryl oxygen.
inely believe it will displace a significant part of current technology." The catalyst's origins lie in Atalla's quest for the logical next step beyond biopulping. In 1990, he was in search of a catalyst that would oxidize lignin in wood pulp fast enough to be industrially practical. Unlike the storage of wood chips for biopulping, holding wood pulp for "two weeks in a reactor is unacceptable when you're talking about a mill that's produc ing 1,000 tons of pulp per day." Atalla wanted to find "an inorganic system that would be stable at high temperatures and oxidize lignin in an hour or two." He hy
pothesized that a catalyst modeled after porphyrin-type oxidoreductases might work. Atalla told his story to research chem ist Ira A. Weinstock, who had just come on board at FPL to work on peroxidase enzymes. Shortly thereafter, Weinstock read about inorganic catalysts being de veloped by chemistry professor Craig L. Hill at Emory University, Atlanta, that just might solve the delignification problem. Hill joined in the quest for the sought-af ter catalyst, collaborating with Wein stock in the development of a promising candidate. The POM technology is being develMARCH 23, 1998 C&EN 41
science/technology
Hammel: fungi use 1-electron oxidation
Atalla: frontier polyoxometalate technology
BKi^e-:-.':.
Weinstock: more POM catalysts Hill: self-buffering catalyst is are on the way truly novel
oped by a partnership that includes FPL, that's intrinsically stable thermodynamiEmory University (with support from the cally under all operating conditions— National Science Foundation), the Uni- bleaching, wet oxidation, filtering, you versity of Wisconsin, the Department of name it," says Weinstock. "If you have Energy, and five companies—four in the something that's intrinsically stable, then U.S. and one in Finland. if it gets damaged during operation it will "We can start with pulp that has as tend to re-form itself." much as 10 or 15% lignin," Atalla says. Meanwhile, professor of biochemistry That would reduce the load of lignin that Karl-Erik L. Eriksson at the University of had to be removed in kraft pulping, Georgia, Athens (UGA), has developed which would cut back on the amount of an enzyme-based process for bleaching chemicals needed, he explains. pulp. The EnZone process, now in pilotPulp capacity would increase 35 to plant testing, uses xylanase in combina40% if chemical pulping were used to tion with oxygen, ozone, and hydrogen take lignin content down to 10% and if peroxide. the POM catalyst were used to bring it In the mid-1980s, Liisa Viikari, a prodown to 5%, says Atalla. In fact, the POM fessor of biotechnology in the forest catalyst conceivably could be interjected industry at VTT Biotechnology & Food earlier, and raise capacity even more, he Research, a research institute in Espoo, suggests. Finland, began pretreating pulp with xyla"It's all very rewarding," says Hill. "The companies are excited. Everyone realizes there is something very valuable here. The one thing truly novel is that the catalyst has its own buffering system. It allows us to work in water with no additives and maintain the pH. And that's critical to prevent damage to cellulose." Bleaching with the POM catalyst involves only two steps. In the anaerobic first step, the catalyst partially oxidizes lignin and is reversibly reduced. In the second step, oxygen is added to the bleaching liquor and the catalyst converts the dissolved lignin to carbon dioxide and water. "The two steps use only air and water, an ideal process that only nature has achieved to date," says Hill. The researchers are still developing new polyoxometalates. FPL chemical engineer Freya Tan examines pulp "We're trying to fashion a catalyst from deinked paper. 42 MARCH 23, 1998 C&EN
nase as a way to cut down on the amount of chlorine needed in bleaching, says Eriksson. Xylanases attack hemicelluose, loosening the polysaccharide's covalent grip on lignin and leaving the exposed lignin more vulnerable to chlorine oxidation. The first industrial trials to use the xylanase approach were conducted in Finland in 1988. Since then, the approach has been widely adopted in Europe and North America, he notes. Eriksson's EnZone process avoids the use of chorine altogether, producing pulp with a very high degree of brightness—a measure of bleaching efficiency. Currently, he is running trial experiments using a xylanase made by the hyperthermophilic bacterium Thermotoga maritima. The enzyme operates at 92 °C throughout the 90-minute bleaching stage and loses only 20% of its activity after 24 hours at that temperature, he says. The enzyme has been cloned into Escherichia colt, but "needs a better production system," he explains. To increase the expression of the enzyme, his team is cloning it in a different organism. With coworker Jan L. Yang, Eriksson has developed an enzymatic deinking process, Enzynk, that uses cellulases to deink recycled office paper, newsprint, and magazines. Conventional deinking "beats the ink off the paper," he says, and uses up a lot of mechanical energy. In 1994, the two scientists founded a company—Enzyme Deinking Technologies (EDT), in Atlanta—to market their new process. The market is growing fast as more and more pulp and paper mills in the U.S., Europe, and Asia begin to adopt the technology, says Eriksson. Enzynk is not an all-purpose deinker, however. "Every mixture of recycled paper requires its own enzyme mixture," he points out. EDT determines
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seience/teehnology Ian R. Hardin, head of the university's textile science department. The alliance is establishing an infrastructure for biotechnology, telecommunications, and environmental programs statewide. It includes a consortium of universities and industry partners, Hardin says. Among ongoing biotechnology efforts is the search for an enzyme that will free flax fibers from the plant's stalk, a process known in the linen industry as retting. The effort is spearheaded by Eriksson and microbiologist Danny E. Akin at the USDA Agricultural Research Service's Richard B. Russell Agricultural Research Center, Athens. Collaborators include researchers in UGA's textile science program; Clemson University, Clemson, S.C.; and Agricultural Research Service facilities in Clemson and New Orleans. Eriksson: bringing biotechnology to Georgia "There's no linen industry in the U.S.," says Akin, although relics of a few which blend will work best for a partic- flax-processing mills remain. Flax was ular application and carries out full- grown for many years in South Carolina scale experiments before commercializ- as a source of cigarette paper, he explains. Now the state is looking to proing the product. "I came to Georgia [from the Swedish duce it for use in textiles. Pulp & Paper Research Institute] to de"We don't intend to start a traditional velop biotechnology for the pulp and pa- linen industry in the U.S.," he says. The per industry in the southeastern U.S. idea is to find an economical route to obwith a special focus on Georgia," says taining short-fiber flax "that could be Eriksson. His position as an eminent blended to give cotton value-added propscholar was set up by the University of erties. It's a new paradigm. The flax would be processed using cotton equipGeorgia Board of Regents. Funding for biotechnology research ment, rather than expensive equipment and development at UGA is boosted by manufactured solely for flax." But new the state-established Consortium on retting methods also would benefit the Competitiveness for the Apparel, Carpet traditional linen industry, he notes. & Textile Industry as well as by the GeorA consortium of state, federal, and ingia Research Alliance, established by dustry groups are working to promote Governor Zell Miller in 1990. This year the development of a flax industry in the alliance's budget is $42 million, says the U.S., says Akin. To that end, they aim to establish a nonprofit industry—the Center for American Flax Fiber— at Clemson University "in the near future," he says. Retting is the first step in the arduous road from flax to linen. Historically, harvested flax was soaked in ponds or lakes where "anaerobic bacteria degraded the glues and pectins that held the fibers together," Akin explains. Environmental laws have put an end to that approach, he says, because of the stench University of Georgia's pilot-plant manager Ryan B. Adolphson, an agricultural engineer, uses a converted milk and pollution produced truck to deliver wood pulp to the plant for bleaching. by the fermentation. 44 MARCH 23, 1998 C&EN
Scanning electron micrograph of flax being retted enzymatically shows fibers (center) partially detached from the waxy sheath (top) and Hgnified core cells (bottom).
Now flax is "dew-retted." First, the plant is pulled from the ground and laid out on the field. Then, when moisture conditions and the temperature are right, indigenous fungi colonize the plant and begin to degrade it. Waiting for that to happen can take from four to eight weeks, Akin says. Overretting degrades cellulose, so "you need to be able to judge when the right degree of retting is reached," he notes. "And dew-retting takes up quite a bit of land." Optimum dew-retting conditions are found in northern France and in Belgium, he says. After retting comes scutching, a beating that separates flax fibers from the woody inner core material. (Flax's long bast fibers run longitudinally just inside the "bark" of the plant.) Scutched flax is hackled (combed) to provide the straight long-line fibers used in spinning traditional linen, Akin says. The Georgia approach would sidestep the scutching and hackling procedures. In preliminary work, Akin and his coworkers have retted flax and spun fibers on a laboratory scale. They first chopped up the flax to give them the short fibers they're after, then retted them with a mix of commercially available pectinases and a calcium chelator that former postdoctoral fellow Gunnar Henriksson (now back in Sweden) showed could help break down pectin.
Bioprospectors with Georgia on their minds Unlike the bioprospectors who stalk their bacterial prey in deep-sea hydrothermal vents or in freezing arctic waters, a group of researchers at the University of Georgia, Athens, are foraging for microorganisms right in their own backyards. Some are looking for bugs that will degrade nylon and stop the need for landfilling carpet waste. Others are after anaerobic fungi that will digest fibrous plant material and pave the way for a new era of biotechnology in the cotton, naturalfibers,and other industries. "We have successfully composted wool with indigenous microbes and turned it into 'fertilizer,' " says associate professor of textile science Patricia A. Annis. "We hope to commercialize that technology this year." The university's department of biological and agricultural engineering collaborated in the woolcomposting project, she notes. Next on the composting list is nylon, which, like wool, is a polyamide. That's enough to make the Georgia team believe that somewhere there's a microbe that can be fooled into feasting on nylon. There's a real need to develop a way to biodegrade carpet waste in Georgia. The state manufactures roughly 60% of the carpets produced in the U.S., which is 40% of the global output, explains professor of biochemistry Karl-Erik L. Eriksson. "There's a lot of carpet waste that goes to landfills. But landfilling is getting very expensive, and available land is getting scarce, so we're looking for microorganisms" that will do the job.
Once a microbe is found, its nylondegrading enzyme can be cloned, Eriksson says. Ideally, he would like to lay his hands on an enzyme that breaks nylon into its monomeric units, which could be recycled. For Lars G. Ljungdahl, director of the university-based Georgia Biotechnology Center, anaerobic fungi are the gold at the end of the rainbow. So far he's isolated 14 different cellulases, xylanases, and esterases from anaerobic fungi that inhabit the rumen of cows—the section of bovine stomach where the first round of digestion takes place. Anaerobic fungi in animals were first discovered in 1975 in sheep, he says. "Think about it," says Ljungdahl. "Where is the best breakdown of plant tissue occurring? It really is in herbivorous animals." Bovine fungi capable of digesting the barrier polymers that shield cellulose in plants could be stashed with enzymes whose job is to break through the obstructing polymers, he says. Such enzymes would be useful in the cotton and wood-pulp and paper industries, which currently depend on energy-intensive or environmentally unfriendly methods to do the job. The fungal enzymes would have applications in the brewing, food, and beverage industries. For example, says Ljungdahl, they could be used to remove pectin for clarifying fruit juices. Anaerobic fungi that make a beeline for cellulose and efficiently siphon off the glucose it's composed of would offer a way to convert plant materials directly
Ljungdahl: exploring bovine fungi
Annis: eyes nylon-degrading fungi
into fuel alcohol, Ljungdahl says. "The Department of Energy is very interested in this." These fungi, like some anaerobic bacteria, degrade cellulose by a mechanism that's not yet fully understood, he explains. On their surface, they carry multiple cellulases anchored in sequence to a long "scaffolding" peptide. Working like a chain gang digging a ditch, the cellulases nick the cellulose chain at regular intervals. After they clip the substrate's glucose units apart, they deliver the pieces directly to the microorganism. The satiated microorganism departs and forms spores, while the network of cellulases (dubbed cellulosomes and discovered in 1975 by Israeli scientists) continues to demolish the cellulose. "The enzymes sit on the cellulose and feed other microorganisms that can't break it down," says Ljungdahl. "This is a system aerobic bacteria don't have," he notes. The enzymes of anaerobic fungi are 10 to 100 times more active than their aerobic counterparts, he says. Last year, Ljungdahl started a company (Aureozyme, in Atlanta) that aims to produce and commercialize fungal enzymes. Currently, he is working to identify and clone the genes that encode the enzymes he has isolated. Georgia's future in biotechnology will be in its textile, wood-pulp, and paper industries, says Ljungdahl. "The state doesn't have a big pharmaceutical industry, but it's coming along pretty well in both agricultural and medical biotechnology." He credits the Georgia Research Alliance (a collaborative effort involving the state of Georgia, six universities, and industry) with helping to fuel biotechnology research. "In the past four orfiveyears, I think we have put about $40 million into biotechnology research in Georgia." Most of the money goes to equipment, eminent scholar support, facilities, and projects that involve both academic and industrial researchers. For example, Ljungdahl notes that X-ray crystallography centers have been established at the University of Georgia and at Emory University in Atlanta. Nuclear magnetic resonance centers also were established at the University of Georgia, Emory, and Georgia State University, Atlanta. "I have never seen such enormous growth and such foresight in support within a university system," Ljungdahl says. "I think this effort by the state of Georgia will have an enormous effect on the future of biotechnology."
MARCH 23, 1998 C&EN 45
science/technology
At left, Akin holds flax that will be cut into short pieces for enzymatic retting. Above, fiber from laboratory-retted flax.
What the group is after, though, is a retting enzyme they can clone. "We have identified the fungus responsible for dew-retting," says Eriksson. "It produces large quantities of endopolygalacturonases." This particular enzyme is important, he says, because it will attack pectin. The fungus produces no cellulases, which is key to preserving the flax fiber, he adds. "We have grown the fungus on a 100-L scale and we are just about to purify the endopolygalacturonases. We will see if this is all it takes to get perfectly retted flax. The question is whether traces of other enzymes also are important. " Using enzymes for retting flax "would improve the cleanliness first of all," because flax won't need to be colonized with fungi, says Akin. "It would also improve consistency, and we hope it can improve quality. Industry can make use of material with low quality, and it can make use of material with high quality. But inconsistent quality drives industry nuts." Some enzymes already are used in textile processing. Amylase is a very old staple in the cotton industry. Since 1857, it has been used to remove starch from woven fabric, says J. Nolan Etters, professor of textile science at UGA. Cellulases took over the stonewashing of jeans in the mid1980s, replacing the rather crude ap46 MARCH 23, 1998 C&EN
proach of pummeling jeans with pumice stones in giant-sized laundry machines to decompose indigo in splotches. That approach was hard on both the fabric and the machinery and "environmentally nasty because it used potassium permanganate and hypochlorite to soak the stones," Etters says. Oxidoreductases are gaining a toehold in stonewashing, he notes, because they don't weaken the fabric as much as cellulases. Current research on cotton is focused on developing enzymatic approaches that will substitute for "scouring" and cut back on the copious washing needed to ready the fabric for dyeing. Scouring is just one of many steps in the transformation of cotton from boll to cloth. The cotton fiber is a single long cell that originates in the seed coat. Thus
Hardin: using enzymes to "scour" cotton
ŒO:
when harvested cotton is ginned to separate the fiber from its seed, not all of the seed-coat fragments are removed. Called neps or motes, they're dealt with later on. Ginned cotton is made into yarn that is woven or knitted into fabric. The yarn is strengthened with a coating of starch, or "sized," to help it withstand the stress of the weaving process. The starch is later removed by treating the fabric with amylase. All of these steps are chemically and environmentally benign. Then comes the not-so-benign scouring, which prepares the woven fabric for the dye bath by removing the fiber's water-insoluble waxy outer layer and pectin shield so dye can penetrate. Sodium hydroxide is used to do the job. "Everyone that makes cotton fabrics scours," says Hardin. "So if you develop an alternative process that would not be putting out the [ionic] salt flow and could work at a lower temperature, you could fundamentally change the cotton industry worldwide." Hardin has planted the seeds for such a revolution. Using a mixture of commercially available pectinases and cellulases, he has developed a way to clip the fiber's pectinaceous material "away from the body of cellulose." The approach has the potential for replacing sodium hydroxide in scouring. But it doesn't take care of the neps and motes. In the traditional approach, these fragments "tend to swell in sodium hydroxide and disappear in the hydrogen peroxide bleaching step that follows," he notes. To help find an enzymatic solution to the seed-fragment problem, Novo Nordisk Biochem, Franklin, N.C., a subsidiary of Denmarkbased Novo Nordisk, is set to team up with Hardin. Separately, the company is developing a catalase that will break down hydrogen peroxide to oxygen and water. "You have to get rid of all traces of hydrogen peroxide before dyeing the fabric," says Brian D. Condon, group leader of enzyme development and applications at the Franklin facility. "Trace amounts can both damage the dye and basically ruin the cloth." What the industry does now, he says, is wash the fabric and rinse it with "huge amounts of water." Novo's catalase would cut down on the water load. Other problems remain. Hardin notes that it's no easy task to get dye into the cotton fiber. Dye molecules tend to move in and out of the fibers, he says, so sodium chloride is added to the dye bath
. . · A must for beginners in SFC and for those to change the ionic strength and help fasten them to the fiber: "Sometimes as much as a pound of sodium chloride per pound of cotton," he says. "A lot of companies are spending a lot of money recovering chemicals used in processing. I think the textile industry believes the goal should be closed-loop processing." Among areas that are ripe for further development, Hardin suggests, is the use of laccases—peroxidases that break aromatic rings—in fragmenting and decolorizing dye molecules in wastewater. New ground was broken this year in permanent-press processing, Hardin notes. UGA's Charles Q. Yang, a professor of textile science, invented a more environmentally friendly way to help keep cotton— and linen—from wrinkling. Both materials wrinkle because their cellulose chains are joined together by hydrogen bonds rather than by covalent cross-links, explains Hardin. When the materials are deformed, the hydrogen bonds are disrupted and re-form in a different position. That's why the fabric must be steam-pressed to heat the molecules, break up the offending bonds, and force them back in a new position, he says. In an upcoming issue of Textile Research Journal, Yang reports a method he has developed to covalently cross-link the cellulose using polymaleic acid and citric acid. It offers an alternative to formaldehyde-based covalent cross-linking methods currently in use. Callaway Chemical Co., Columbus, Ga., already has signed a license agreement with the university to commercialize Yang's process, Hardin notes. Thus, research is moving forward in the textile industry. Hardin says he has "more ideas for research and less time to do it than ever before." Etters believes enzymes have the potential to open a "brand-new area" in coloring. "Look at all the bright colors in blooms in nature. It's all done enzymatically. Suppose you could get colors like that without using harsh chemicals? Wouldn't it be wonderful?" Nevertheless, Hardin, Etters, and other textile researchers realize that no matter how "green" or economical a new process is, it will be a hard sell to industry if it involves any retrofitting. Just as in the pulp and paper industry, the problem is money. As Emory's Hill puts it: "Capital costs in pulp and paper mills are astounding. Even if we have a better mouse trap, we will have to convince people to use it."^
seeking a better, faster, and safer way to analyze
data...
American Chemical Society Presents A New Lecture-Laboratory Short Course
Supercritical Fluid Chromatography Monday-Friday, May 11-15, 1998 Virginia Tech · Blacksburg, Virginia This Intensive, Five-Day Course Will Teach You: • How to interpret SFC data in terms of retention, reproductibility, and repeatability • Which strategies to use when your analyte will not dissolve in C0 2 • Why and when SFC offers advantages over H PLC and GC • When open tubular and packed columns should be used • How to work with multiple SFC instruments • The extent that all detection devices can be successfully interfaced to SFC • And much more!
Course Instructors Larry T . Taylor, Virginia Tech, Blacksburg, VA Thomas L. Chester, Procter & Gamble, Cincinnati, OH Terry A. Berger, Berger Instruments, Newark, DE Register Today—Registrations are limited! Call the ACS Short Course at (800) 227-5558, ext. 4508 (TOLL FREE) or (202) 872-4508. For more information, mail or fax the coupon below.
^ _ .
fySS!Please send me a brochure describing the new ACS Short Course, Supercritical Fluid Chromatography, to be taught May 11-15,1998, at Virginia Tech in Blacksburg, VA.
Name_ Title. Organization _ Address City, State, Zip_ Phone Fax — E-mail _
Mail to:| A m e r i c a n Chemical Society, Dept. of Continuing Education, Meeting I Code VPI9805, 1155 Sixteenth Street, N . W . , Washington, D C 20036. F A X . (202) 872-6336. E-mail:
[email protected] MARCH 23, 1998 C&EN
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