Compared with first-generation U.S. ethanol production currently centralized in the Corn Belt, the emerging second-generation industry will exhibit much greater diversity in location, feedstock and process. Feedstock supply and geographic location are naturally tied to each other, and this is displayed in all of the plants dotting the Midwest. There is a subtler but definite connection between process design and location.

Standardizing a process design is much more calculable when every project is designed to use the same feedstock. For those processes based on locally or regionally available feedstock supplies, which include wood, stover, straw, dedicated energy crops or hundreds of other lignocellulosics, standardizing any one process for a myriad of potential feedstocks has proven to be more difficult. This is why so many viable processes have emerged in second-generation ethanol project development, including thermochemical, biochemical and hybrid technologies.

For biochemical processing, enzymatic hydrolysis can play an important role in overall process design but even within this specified aspect of biochemical conversion, there is no clear-cut image of what the second-generation enzyme manufacturing and supply chain will look like. Those closest to the situation tell EPM there are still a lot of factors that will influence how this particular aspect of a second-generation ethanol industry is likely to unfold. Moving forward, however, it seems evident enzyme manufacturing and delivery will be as varied as project location, feedstock choice and process design.

In corn-ethanol production, alpha amylases help break down complex starch carbohydrates into polysaccharides during liquefaction. Then, during saccharification, glucoamylases further deconstruct polysaccharides into simple glucoses ready for fermentation.

For first-generation enzymes, centralized production by major enzyme-manufacturers is the norm, followed by transportation and delivery to the various dry-grind ethanol plants. Novozymes has one enzyme manufacturing plant in the United States, located in Franklinton, N.C., from which it serves U.S. customers as far away as the West Coast. According to Emmanuel Petiot, Novozymes global business development manager, this centralized model has been successful thus far in first-generation ethanol enzyme production and delivery, but times are changing. "Yes this model has been one that has worked but, because of the increased size of the first-generation ethanol industry, in addition to the volume of enzymes needed for biomass [hydrolysis], we need to get closer to our customers," he says. "That's a given."

The Case for On-Site or Near-Site Enzyme Manufacturing
Novozymes' first step in developing what Petiot coins as its near-site model for enzyme production, as compared with its current strategy of centralized manufacturing, is its new enzyme plant project in Blair, Neb. Construction on the enzyme plant is slated to begin late this year, with operations expected to start in late 2010.

"Initially, the [Blair] plant will primarily serve first-generation plants," Petiot says. "But as pilot, demonstration and the first commercial biomass ethanol facilities come on line, we'll serve these plants as well. The number of biomass plants will be small in the beginning—maybe six, eight or 12 of them by 2012—but it will put us in the position to serve all of the needs of the biomass industry in its first years of development."

In order to deconstruct and hydrolyze the polymeric sugars imbedded in extremely complex structures of lignocellulosic materials, cocktails of cellulases and hemicellulases are required. "The volumes of enzymes required for biomass are going to be much higher than for first-generation ethanol because the feedstocks are much more complex," Petiot says.

Verenium Corp. vice president of research and development, Nelson Barton, agrees. "We are talking very large volumes of enzymes," Barton tells EPM. "The nature of the lignocellulosic substrate will require more complex enzyme cocktails and, when you start talking about manufacturing very large volumes of enzymes, you're talking about transporting large volumes of water around as well."

In 2007, Verenium was formed through the merger of Cellunol (formerly BC International) and enzyme-maker Diversa Corp., putting Verenium in the unique position of possessing its own specialty enzymes division. "Out of our Diversa heritage comes our development of process-optimized enzymes," Barton says. "We do plan to produce a targeted set of [second-generation] enzymes, but we also have the ability to optimize and evolve these enzymes and make them more stable."

Within minimal processing and minimal stabilization models, Verenium is working to process-optimize and stabilize its cellulase and hemicellulase cocktails for biomass hydrolysis. "The farther you need to transport the enzymes, the more you need to start thinking about stability issues during transport," Barton says. Issues of cost are huge in all aspects of the emerging biomass industry, so if second-generation enzyme cocktails are transported long distances to a biorefinery and the enzymes destabilize en route, how much processing of the enzymes can you afford to do after you've produced them in order to stabilize them? Barton furthers this question by asking, "Could you even afford to do any sort of formulations to stabilize them in any way?" He responds to his own question by stating, "The answer is minimal, if any. So really it becomes a matter of being much more amenable to on-site manufacturing of enzymes."

Clearly the enzyme manufacturing and logistics chains will be evolving over time. "Early on it's going to be on-site production until the industry gets to a point where regional manufacturing makes sense," Barton says. Verenium plans to supply its own cellulosic ethanol plants with enzymes—the first of its demonstration plants, the 1.4 MMgy facility in Jennings, La., is up and running—and Barton says the company is willing to supply enzymes through regional distribution or on-site production as needed. "If [non-Verenium] plants were to require on-site enzyme production in order to meet their logistics needs, then that is something we can do," Barton says.

Verenium and its specialty enzymes division is not the only player advocating enzyme production on-site of a biorefinery complex. Genencor International's vice president of biorefineries, Jack Huttner, tells EPM there are several well-known challenges to using cellulases for commercial-scale conversion of biomass to ethanol—including the economic and technical challenges of transporting enzyme cocktails.

"In general, cost savings through on-site enzyme production would be significant considering shipping and storage costs, as well as the potential to share energy infrastructure costs with the main ethanol plant," Huttner says.

Genencor, a division of Danisco A/S, has what Huttner characterizes as a two-pronged approach to meeting the many challenges facing a biomass-based ethanol industry. One of these "prongs" is called DuPont Danisco Cellulosic Ethanol LLC, a joint venture between DuPont and Genencor. The joint venture's latest development is a pilot plant and process development project with the University of Tennessee's Genera Energy LLC, which is focused on two specific feedstocks: corncobs and switchgrass. "DuPont Danisco Cellulosic Ethanol is on a fast-track to develop and license an integrated biorefinery package," Huttner says. This biorefinery package covers the entire production process from feedstock collection, to pretreatment and hydrolysis, to fermentation and recovery, and will include on-site enzyme production for the licensee. "Producing enzymes at the ethanol production site would allow for numerous advantages in production time, cost and energy efficiencies, as well as equipment and material quality control," he says.

Genencor's second avenue to supply enzyme cocktails for the biomass industry is through its more conventional-style merchant enzyme business using a centralized distribution system more resembling first-generation logistics. However, even then Huttner says, "Genencor is willing to discuss on-site enzyme production with potential beorefinery operators, or enzyme supply from one of Genencor's regional manufacturing plants."

Along with the higher volumes of enzymes needed to hydrolyze biomass and shipping these formulated products there's another factor to consider, according to Verenium spokeswoman Morgen Grandjean. "It's also a matter of carbon footprint," she says. Transporting huge volumes of enzymes entails moving large quantities of water to keep the enzymes stable and active. Grandjean stresses that on-site enzyme production would help keep the carbon footprint of the enzyme industry to a minimum.

The Case Against On-Site Enzyme Manufacturing
Petiot admits the topic of making cellulases and hemicellulases on-site of a biorefinery is chic today. "This is something everyone is talking about," he tells EPM. However, he says Novozymes has not yet reached the conclusion that this is going to be the case—at least for Novozymes. "We believe that manufacturing enzymes is a complex technology," he says. Because of the complexities involved in the manufacturing of enzyme mixes for a biomass-based ethanol industry, Petiot says it could prove to be difficult to produce everything needed on-site. "Say if your feedstock changes, or if enzyme suppliers find new and more efficient activities, it is not easy to switch from one production strain to another," he explains. In other words, on a technical basis alone, there may be limits to what can be done with enzyme manufacturing at a biorefinery.

First-generation ethanol production from corn requires alpha and gluco amylases, but second-generation hydrolysis of lignocellulosic materials will likely require a minimum of four enzymes: endogluconase, cellobiohydrolase 1, cellobiohydrolase 2 and beta glucosidase for starters. Cocktail formulation depends not entirely on feedstock selection but also on which pretreatment method is incorporated into the overall process design. An enzyme cocktail for a producer using a mild acid pretreatment, which converts the hemicellulose to monomeric sugars, is going to be formulated differently than an enzyme mix for a producer using a more alkaline pretreatment. "An alkaline pretreatment process would require you to add [hemicellulases] to convert hemicellulose to C5 sugars, in which case you're talking about adding xylanases and hemicellulases to complete that cocktail," Barton says. "It comes back to having your core workhorse enzymes that do a lot of work on the primary cellulose component and, depending on the feedstock preferences and region, the slightly different compositions of your hemicellulose fractions. I could see the need to optimize the basic cocktails by adding accessory enzymes to optimize the performance on a given substrate."

A biorefinery based on the biochemical process platform using enzymatic hydrolysis may have multiple feedstock supply arrangements, which again would require switching to slightly different formulations of proteins and activities. "That is not going to be easy for an on-site production facility to do, whereas in a more centralized site you can switch around these activities more easily because enzyme formulation is all that is done at that location," Petiot says. "If the industry picks up as we expect, and if we have to build several [near-site] facilities, then those enzyme manufacturing sites will be able to manufacture these complex activities, but also be able to switch them around in a more efficient manner."

Ron Kotrba is an Ethanol Producer Magazine senior writer. Reach him at rkotrba@bbibiinternational.com or (701) 738-4942.