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Research Summary: Professor Lee Lynd
Introduction
My group seeks to be of service by enabling and envisioning a transition from the non-sustainable present to a sustainable future, one of the primary challenges facing humanity in the twenty-first century. Our particular focus is on utilization of plant biomass, a central and essential intermediate in a world supported by sustainable resources. As presented in Figure 1, most paths from sustainable resources to human needs pass through either renewable electricity or biomass, with biomass being the sole foreseeable sustainable source of organic fuels, chemicals, and materials as well as food.
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| Figure 1. Sustainable resource supply and utilization |
Among various forms of plant matter, cellulosic biomass - that is, biomass composed primarily of inedible plant fibers having cellulose as a prominent component - has the largest potential contribution in the context of sustainable and secure provision of energy services due to economic, scale of supply, and environmental considerations (17,27).
As represented in Figure 2, my group approaches utilization of cellulosic biomass as a sustainable resource from three intellectual vantage points - applied biology, process engineering, and analysis of resource and environmental efficacy - addressed below.
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| Figure 2. Lynd group activity and motivation map |
In the course of bringing these diverse intellectual approaches to bear on fostering the sustainable resource transition and advancing biomass conversion and utilization, we frequently encounter frontiers of fundamental understanding. For example, evaluating the feasibility of microbial conversion of cellulosic biomass without added cellulolytic enzymes (consolidated bioprocessing) is enlightened in important ways by new understanding of the fundamental physiology of cellulolytic microorganisms. Similarly, estimating the performance and cost of a reactor in which cellulose hydrolysis occurs is enlightened by new understanding of the kinetics and multi-phase transport phenomena operative in that reaction. Consistent with the "Pasteur's Quadrant" model articulated by Donald Stokes (Brookings Institution Press, Washington, DC, 1997), we see advancing applied capability and increased fundamental understanding as having strong potential to be convergent and mutually-reinforcing, and we aspire to achieve this potential.
The group's organization and strategy provide unusual opportunities for students and other members to gain exposure, and potentially make contributions, at several levels. These include advancement of fundamental scientific understanding in selected areas, design and evaluation of biological and chemical processing technology, and consideration of "big picture" issues associated with the sustainable resource transition and the role of biomass in this context. The Lynd group is interdisciplinary, in terms of both prior background and current activities, reflecting the opportunities and challenges inherent in our area of focus.
Applied Biology
My group's applied biological research focuses on two related themes: organism development for consolidated bioprocessing and the fundamentals of microbial cellulose utilization. In the course of pursuing these two themes, we are also active in development of laboratory methods that enable us to pursue our goals.
Consolidated bioprocessing. Consolidated bioprocessing (CBP) is a processing strategy for cellulosic biomass which involves consolidating into a single process step four biologically-mediated events: cellulase production, cellulose hydrolysis, hexose fermentation, and pentose fermentation (11,17,25,41). Implementing this strategy requires development of microorganisms that both utilize cellulose and other biomass components while also producing a product of interest at sufficiently high yield and concentrations. Development of such organisms is a potential breakthrough that would result in very large cost reductions as compared to the more conventional approach of producing saccharolytic enzymes in a dedicated process step (10,41). We are focused on production of ethanol, a promising renewable fuel (11,42), via CBP. The CBP strategy is however potentially applicable to a very broad range of products (17), and our progress has significant implications in this context. The feasibility of CBP is supported by kinetic and bioenergetic analysis (16) as well as work undertaken as part of our investigation of microbial cellulose utilization.
Organism development via the native cellulolytic strategy. One approach to organism development for CBP begins with organisms that naturally utilize cellulose and other biomass components, and uses genetic engineering to enhance product yield and tolerance. We are studying Clostridium thermocellum, a thermophilic bacterium that has among the highest rates of cellulose utilization reported, as well as the xylose-utilizing thermophiles Thermoanaerobacterium saccharolyticum and Thermoanaerobacterium thermosaccharolyticum. We seek to eliminate production of acetic and lactic acid in these organisms via metabolic engineering. In addition to increasing the yield of ethanol, our results suggest that organic acid production may be responsible for the low concentrations of produced ethanol generally associated with these organisms (9,19). We have devoted a substantial effort to developing gene transfer systems for the above-described target organisms, and have recently enjoyed substantial success in this area (see Development of laboratory methods), and have also characterized multiple C. thermocellum isolates from nature (24). With these tools in hand, we are actively pursuing metabolic engineering of thermophilic, saccharolytic bacteria. We have recently reported knockout of lactate dehydrogenase in T. saccharolyticum (31), and will soon submit a paper reporting knockout of acetate kinase and phosphotransacetylase in this organism. Mascoma Corp. has licensed technology involving development of thermophilic ethanol-producing microorganisms initiated in the Lynd lab and is working to further advance this approach with continued involvement of the Lynd group.
Organism development via the recombinant cellulolytic strategy. An alternative approach to organism development for CBP involves conferring the ability to grow on cellulose to microorgnanisms that naturally have high product yield and tolerance via expression of a heterologous cellulase system and perhaps other features. My group works closely in this area with Professor Willem van Zyl and his group of the University of Stellenbosch (South Africa). The Stellenbosch group has engineered Saccharomyces cerevisiae to express over two dozen different saccharolytic enzymes (e.g. references in 25). Our role in this collaboration is to characterize recombinant strains with respect to enzyme production (in preparation) and their ability to utilize non-native substrates (38), and also to investigate selection in continuous culture as a means to develop improved strains and/or enzymes (37).
Fundamentals of microbial cellulose utilization. Whereas cellulose hydrolysis has been approached in the literature primarily in the context of an enzymatically-oriented intellectual paradigm, the CBP processing strategy requires that cellulose hydrolysis be viewed in terms of a microbial paradigm. This microbial paradigm naturally leads to an emphasis on different fundamental issues, organisms, cellulase systems, and applied milestones compared to those of the enzymatic paradigm (25). We are working with C. thermocellum as a model organism because of its high growth rate on cellulose together with its potential utility for CBP.
A prominent focus of our effort in this area involves the bioenergetics of microbial cellulose utilization. We have proposed (22,25) a bioenergetic model in which the overall rate of ATP production due to glycolysis, acetate kinase, and phoshorolytic cleavage of β-glucans is equal to the overall rate of ATP consumption due to substrate transport, cell growth, cellulase synthesis, and maintenance. Recently, we showed that the rate of phosphorolytic cleavage of cellodextrins greatly exceeds that of hydrolytic cleavage in C. thermocellum cell extracts (34), confirming the possibility of energy conservation from cleavage of β-glucosidic bonds as suggested by previous studies in the literature. Incorporating this result into our bionergetic model in conjunction with data from continuous culture of cellodextins of various lengths provides strong, albeit indirect, evidence that cellulose utilization by growing cultures of C. thermocellum involves assimilation of cellodextrins with mean length ~ 4 (39). This interpretation was cooroborated (39) by showing that the mixture of cellodextins in cells grown on 14C-cellulose has an average length of 4 with cellopentaose (G5) present at the highest concentrations, whereas cells grown on 14C-cellobiose have an average length of 2 with cellobiose present at the highest concentrations. Taken together, our recent results indicate that 1) C. thermocellum hydrolyzes cellulose by a different mode of action from the classical mechanism involving solubilization by cellobiohydrolase, 2) bioenergetic benefits specific to growth on cellulose are realized resulting from the efficiency of oligosaccharide uptake combined with intracellular phosphorolytic cleavage of β-glucosidic bonds, and 3) these benefits exceed the bioenergetic cost of cellulase synthesis, supporting the feasibility of anaerobic biotechnological processing of cellulosic biomass without added saccharolytic enzymes.
A recent manuscript (35) presents data on control of cellulase synthesis in batch and continuous cultures of C. thermocellum, and presents evidence that cabolite repression is operative in this organism. In addition, our data suggests that either growth substrates in addition to cellobiose are taken up during cellulose utilization by growing cultures and/or that the presence of insoluble cellulose triggers an increase in cellulase synthesis. Additional work in the area of microbial cellulose utilization targets quantitative evaluation of enzyme-microbe synergy and examination of the degree to which hydrolysis products equilibrate with the bulk solution. Recent reviews address understanding and modeling of cellulose hydrolysis by non-complexed cellulase systems (36), as well as the fundamentals of microbial cellulose utilization by C. thermocellum (43).
Development of laboratory methods
Gene transfer systems. We have developed the first gene transfer system for C. thermocellum (33) based on novel electroporation (ET) apparatus developed by Visiting Professor Mikhail Tyurin (7). A recent manuscript (44) examines the role of current oscillations in achieving high-efficiency ET, and also reports efficient transformation of Thermoanaerobacterium saccharolyticum. These techniques enable metabolic engineering of thermophilic bacteria pursuant to consolidated bioprocessing. Earlier work characterized restriction systems in C. thermocellum (12,21) and T. saccharolyticum (12). Recently we have reviewed gene transfer protocols for thermophilic bacteria in a chapter appearing in Methods in Microbiology (45).
Continuous culture on cellulosic substrates. The Lynd lab has developed custom laboratory systems for non-batch microbial cultivation featuring metered, asceptic addition of particulate cellulosic feedstocks. These systems have been used to investigate cellulose utilization by C. thermocellum (1,3,35) as well as simultaneous saccharification by Saccharomyces cerevisiae in conjunction with an added cellulase preparation (5,26).
Cellodextrin preparation. A method for preparing gram quantities of purified cellodextrins of length 3 through 6 was developed using mixed acid hydrolysis followed by chromatographic separation (29). Substrates prepared using this method were instrumental in our recent study of the bioenergetics of cellulose utilization by C. thermocellum (39), and are currently being used in several related studies.
Independent quantification of cell and cellulase concentrations. A method making use of enzyme-linked immunosorbent assay (ELISA) was developed that allows the concentration of cells and cellulase to be determined in cellulose-grown cultures of C. thermocellum (22,30). This method has been used thus far in studies of bioenergetics (39) and control of cellulase synthesis (35).
Cellodextrin phosphorylase assay. To measure the rate of reaction catalyzed by cellodextrin phosphorylase (CdP) in the chain-shortening direction at elevated temperatures, a protocol was developed featuring discrete sampling at 60 degrees C followed by subsequent analysis of reaction products (glucose and glucose-1-phosphate) at 35 degrees C (34). This method was used to evaluate the relative importance of phosphorolytic and hydrolytic cleavage of cellodextrins by C. thermocellum as well as for characterization of CdP kinetics (34).
Improved method for determination of cellulose degree of polymerization. The number average degree of polymerization was calculated as the ratio of glucosyl monomer concentration using the phenol-sulfuric acid to reducing end concentration using a modified 2,2'-bicinchoninate (BCA) method (40). This method is expected to be valuable for studies aimed at understanding the relationship between the physical properties of cellulosic substrates and the rates of biologically-mediated hydrolysis.
Process Engineering
Design and evaluation of advanced biomass conversion processes. Process design and evaluation is an important contribution in its own right while also providing guidance for efforts to advance biomass processing technology through laboratory research. We focus particularly on anticipating the cost and performance of mature biomass processing technology - that is, technology comparable to today's oil refineries for which additional research-driven cost reductions are expected to be incremental. Although admittedly uncertain, features of mature technology are important in the context of evaluating the potential of biomass conversion processes and hence the effort appropriate to expend pursuant to realization of this potential. In 1996 we published an analysis that projected a most likely selling price of 50 cents per gallon for ethanol produced by mature technology (10). We later expressed the results of this analysis in terms of feedstock transfer cost, emphasizing the relevance of mature biomass processing technology to a broad range of products (17). In conjunction with a project entitled "The Role of Biomass in America's Energy Future" (see below), we have collaborated with Eric Larson's group at Princeton in a comprehensive analysis of advanced biomass conversion processes. Results from this analysis have appeared in a report entitled "Growing Energy: How Biofuels can Help End America's Oil Dependence" (PDF) prepared for the National Commission on Energy Policy, with more complete presentations of this analysis to appear in a special issue of Biomass and Bioenergy with supporting materials to be posted on the web.
Reactor design for biological processing of cellulosic biomass. The kinetics of cellulose hydrolysis mediated by both enzymatic and microbial catalysts is complex, very different from kinetics for soluble substrates, and poorly understood. In 1995 (8), we proposed that three features are essential to incorporate into models for enzymatic hydrolysis: adsorption exhibiting saturation in either substrate or enzyme; substrate reactivity that declines with increasing conversion; and, for continuous reactors, a model that considers the changing rate of reaction of particle populations of different ages over the time they spend in the reactor. This model has been expanded to consider differential retention of solids (6). Recently (Shao et al. in preparation), we reformulated this model to simulate intermittent feeding and to eliminate iterative loops. We have also initiated an effort to develop more mechanistically detailed models that consider the action of different cellulase components as well as physical properties of the substrates, have prepared a substantial review of such models (36), and have proposed a substantial new model (46).
The biomass conversion processes we envision will feature bioreactors and other unit operations that involve very large volumes, at least two and often three phases, as well as complex reaction kinetics. Systematic development of understanding and correlations that allow rational scale-up of such processes is in its nascent stages and has clear potential to substantially accelerate commercial deployment. A joint project aimed at addressing this need was initiated by Professor Lynd's and Adjunct Professor Wyman's groups with support by a major grant from the National Institute of Standards and Technology to address this need. We are collaborating with Fluent Inc., a leader in computational fluid dynamics (CFD), to develop state-of-the-art models for multiphase bioreactors based on computational fluid dynamics, and to incorporate kinetic models into this framework for use in scale-up analysis.
Production of ethanol from paper sludge. Paper sludge is a solid by-product of pulping and paper-making operations that is currently disposed of primarily in landfills. We have carried out a comprehensive study showing that many sludges are highly reactive to hydrolysis mediated by cellulase enzymes (18). Paper sludge is an attractive point-of-entry and proving ground for commercial processes featuring enzymatic hydrolysis of cellulose because of potential revenues from avoided sludge disposal, the possibility of additional revenue from recovering mineral components from sludge, no requirement for a pretreatment step to make the substrate accessible to cellulases, and the potential availability at a paper mill of steam, power, and wastewater treatment at incremental cost. In addition, conversion of paper sludge to ethanol and recovered minerals offers a route to nearly eliminating the largest waste stream associated with the pulp and paper industry. We describe developed a unique bioreactor capable of aseptic, metered feeding of solid paper sludge (26). Carrying out simultaneous saccharification and fermentation in this reactor, we show that > 90% hydrolysis yields can be achieved while producing economically-recoverable ethanol concentrations (26). An in-preparation manuscript present evaluation of mixing characteristics and feeding frequency, with a second manuscript presenting process design and economic analysis for paper sludge conversion to ethanol.
Biomass pretreatment. Pretreatment is required to make cellulosic biomass amenable to enzymatic hydrolysis. Working in collaboration with Michael Antal's group at the University of Hawaii, we have published several papers on liquid hot water pretreatment (13,20,23). We showed that liquid hot water pretreatment can, at low solids concentration, achieve performance comparable to dilute acid hydrolysis without some of the complications that use of acid entails. Our results provided a substantial set of data useful for both evaluating liquid hot water pretreatment in comparison to other approaches and for understanding the fundamentals of chemical and physical processes operative during pretreatment.
Analysis of Resource and Environmental Efficacy.
John Prausnitz (1991. Chem.-Ing.-Tech. 63:447-457) wrote: "If engineering is the application of science for human benefit, then the engineer must be a student of not only the application of science, but of human benefit as well." Consistent with this sentiment, I have been active throughout my career in analysis of the resource and environmental efficacy of biomass conversion processes.
The Role of Biomass in America's Energy Future. Launched in the spring of 2003, "The Role of Biomass in America's Energy Future" (RBAEF) project seeks to identify and evaluate paths by which biomass can make a large contribution to meeting future demand for energy services, and to determine what can be done to accelerate biomass energy use and in what timeframe associated benefits can be realized. The project is co-led by Nathanael Greene of the Natural Resources Defense Council (NRDC). Technology-related analysis associated with the "RBAEF" project is sponsored by the Department of Energy Office of Energy Efficiency and Renewable Energy. Environmental and policy analysis is supported by the Energy Foundation and the National Commission on Energy Policy. Participating institutions include - in addition to Dartmouth and the NRDC - Argonne National Laboratory, Michigan State University, the National Renewable Energy Laboratory, Oak Ridge National Laboratory, Princeton University, the Union of Concerned Scientists, the University of Tennessee, and the USDA Agricultural Research Service.
Project tasks focus on: 1) Feedstock production and utilization, focusing on switchgrass as a model cellulosic crop and including the possibility of recovering animal feed protein in conjunction with production of a processing feedstock; 2) Process technology, including biological and thermochemical technologies as well as mobility chain analysis, 3) Analysis of resource sufficiency, focusing on whether and how sufficient biomass could be available to meet very large scale needs such as transportation, 4) transition dynamics, investigating rates of technology improvement and deployment capacity increase under various scenarios, and 5) policy formulation and evaluation. The RBAEF project is unprecedented with respect to the breadth of technologies considered and the diversity of participants involved - representing the technical, environmental advocacy, and policy communities. The project is also differentiated from prior studies by adopting a "high beam" perspective focusing on mature technology that can reasonably be expected in the future given a concerted effort.
Results from this analysis have appeared in a report entitled "Growing Energy: How Biofuels can Help End America's Oil Dependence" (PDF) prepared for the National Commission on Energy Policy, with more complete presentations of this analysis to appear in a special issue of Biomass and Bioenergy with supporting materials to be posted on the web.
Life cycle analysis and resource assessment.
A series of our papers address greenhouse gas emissions and net fossil fuel
displacement on a life-cycle basis (2,4,11,14,15,27), and have contributed
to increasing acceptance of the positive potential of processes based on
cellulosic biomass in terms of these metrics. Most recently, a paper in the Journal
of Industrial Ecology (32) presents a framework that identifies key feedstock,
process, and product parameters that determine net fossil fuel displacement,
(kg fossil fuel displaced/kg product). It is further shown that a broad
range of outcomes, from > 1 to < 0, is possible depending on
the features of the process as represented by values of these parameters. A paper by McLaughlin et al. with coauthors including Professor Lynd (24) examines production of switchgrass from economic and environmental perspectives. An additional paper examines prospects for biomass conversion in South Africa (28).
Biomass energy production and utilization scores well in terms of many metrics evaluated on a normalized basis. Thus for example, favorable values are expected to be realizable for biomass-based energy production in terms of fossil fuel input per ton biomass, soil carbon per acre, greenhouse gas emissions per mile, and dollars per gallon. However, resource issues, related to the number of units of biomass processed, are less well understood and present a more substantive challenge. An in-depth review and analysis of this issue will appear in an invited book chapter (47).
Policy development and evaluation.
Professor Lynd is an active participant in policy analysis related to biomass energy production and utilization. For example, Dr. Lynd has recently played an integral role in policy formulation carried out by the Energy Future Coalition and Role of Biomass in America's Energy Future, and has been invited to present policy-related input to the Governor's Ethanol Coalition, the 25 x '25 group*, the National Commission on Energy Policy, and the Agriculture Committee of the U.S. Senate. In 1994 and 1995, Professor Lynd was the biofuels industry representative on a 30-person committee advisory to the Executive Office of President Clinton on Reducing Greenhouse Gas Emissions from Personal Vehicles. Members of Professor Lynd's research group have opportunities for exposure and involvement in policy-related analysis.
*A group of farm leaders motivated by the goal of farms producing 25% of the energy used in the U.S. by 2025.
References (encompassing many, but not all, Lynd group publications)
47) Lynd, L.R., M. Laser, J. McBride, K. Podkaminer, J. Hannon. Energy myth three - High land requirements and an unfavorable energy balance preclude biomass ethanol from playing a large role in providing energy services. Invited chapter to appear in: B. Sovacool and M. Brown (eds) Energy and Society: Forteen Myths About the Environment, Electricity, Efficiency, and Energy Policy in the United States. Springer.
46) Zhang, Y.-H.P., L.R. Lynd. 2006. A functionally-based model for hydrolysis of cellulose by fungal cellulase. Biotechnology and Bioengineering. 94(5): 888-898. (Abstract)
45) Tyurin, M.V., L.R. Lynd, J. Wiegel. 2006. Gene transfer systems for obligately anaerobic thermophilic bacteria. Invited chapter in: A. Oren and F. Rainey, eds. "Extremophiles", Methods in Microbiology, 35: 309-330.
44) Tyurin, M.V., C.R. Sullivan, L.R. Lynd. 2005. Role of spontaneous current oscillations during high efficiency eletrotransformation of thermophilic anaerobes. Appl. Env, Microbiol. 71:8069-8076. (Abstract)
43) Lynd, L.R., P.J. Weimer, G. Wolfaardt, Y-H.P. Zhang. Cellulose hydrolysis by Clostridium thermocellum: A Microbial Perspective. Invited chapter in: Kataeva I.A., ed. Cellulosome. Nova Science Publishers, Inc., Hauppauge, NY, USA. In press.
42) Greene, N., F.E. Celik, B. Dale, M. Jackson, K. Jayawardhana, H. Jin, E.D. Larson, M. Laser, L. Lynd, D. MacKenzie, J. Mark, J. McBride, S. McLaughlin, D. Saccardi. 2004. Growing energy. How biofuels can help end America's oil dependence. (Article)
41) Lynd, L.R., W.H. van Zyl, J.E. McBride, M. Laser. 2005. Consolidated bioprocessing of cellulosic biomass: an update. Curr. Opin. Biotechnol. 16:577-583. (Abstract)
40) Zhang, Y.-H.P., L.R. Lynd. 2005. Determination of the number average degree of polymerization of cellodextrins and cellulose with application to enzymatic hydrolysis. Biomacromolecules. 6:1510-1515. (Abstract)
39) Zhang, Y.-H.P., L.R. Lynd. 2005. Cellulose utilization by Clostridium thermocellum:bioenergetics and hydrolysis product assimilation. PNAS. 102:7321-7325. (Abstract)
38) McBride, J.E., J. J. Zietsman, W. H. Van Zyl, L.R. Lynd. 2005. Utilization of cellulobiose by recombinant beta-glucosidase-expressing strains of Saccharomyces cerevisiae: Characterization and evaluation of the sufficiency of expression. Enz. Microb. Technol. 37:93-101.
37) Fan, Z., J. E. McBride, W. H. van Zyl, Lee R. Lynd. 2005. Theoretical analysis of selection-based strain improvement for microorganisms with growth dependent upon extracytoplasmic enzymes. Biotechnology and Bioengineering. 92(1): 35-44. (Abstract)
36) Zhang, Y.-H.P, L.R. Lynd. 2004. Toward an aggregated understanding of enzymatic hydrolysis of cellulose: Non-complexed cellulase systems. Biotechnol. Bioeng. 88:797-824. (Abstract)
35) Zhang, Y.-H.P, L.R. Lynd. 2005. Regulation of cellulase synthesis in batch and continuous cultures of Clostridium thermocellum. J. Bacteriol. 187:99-106. (Abstract)
34) Zhang, Y., L.R. Lynd. 2004. Kinetics and relative importance of phosphorolytic and hydrolytic cleavage of cellodextrins and cellobiose in cell extracts of Clostridium thermocellum. Appl. Environ. Microbiol. 70:1563-1569. (Abstract)
33) Tyurin, M., S.G. Desai, L.R. Lynd. 2004. Electrotransformation of Clostridium thermocellum. Appl. Environ. Microbiol. 70:883-890. (Abstract)
32) Lynd, L.R., M.Q. Wang. 2004. A product non-specific framework for evaluating the potential of biomass-based products to displace fossil fuels. J. Indust. Ecol.*** 7:17-32. (Abstract)
31) Desai, S.G., M.L. Guerinot L.R. Lynd. 2004. Cloning of L-lactate dehydrogenase and elimination of lactic acid production via gene knockout in Thermoanaerobacterium saccharolyticum JW/SL-YS485 Appl. Microbiol. Biotechnol. 65:600-605. (Abstract)
30) Zhang, Y, L.R. Lynd. 2003. Quantification of cell and cellulase mass concentrations during anaerobic cellulase fermentation: Development of an Enzyme-Linked Immunosorbent Assay-based method with applications to Clostridium thermocellum batch cultures. Anal. Chem. 75:219-227. (Abstract)
29) Zhang, Y.-H.P., L.R. Lynd. 2003. Cellodextrin preparation by mixed acid hydrolysis and chromatographic separation. Anal. Biochem. 322:225-232. (Abstract)
28) Lynd, L.R., H. van Blottnitz, B. Tait, J. de Boer, I.S. Pretorius, K. Rumbold, W.H. van Zyl. South African J. Sci. Converting plant biomass to fuels and commodity chemicals in South Africa: A third chapter? South African Journal of Science, November/December 2003, 499-507. (Abstract)
27) Lynd, L.R., H. Jin, J.G. Michels, C.E. Wyman, B.E. Dale. 2003. Bioenergy: Background, potential, and policy. Center for Strategic and International Studies. Washington, DC. (PDF File)
26) Fan, Z., C. South, K. Lyford, J. Munsie, P. van Walsum, L.R Lynd. 2003. Conversion of paper sludge to ethanol in a semincontinous solids-fed reactor. Bioproc. Biosystems Eng. 26:93-101. (Abstract)
25) Lynd, L.R., P.J. Weimer, W.H. van Zyl, I.S. Pretorius. 2002. Microbial cellulose utilization: fundamentals and biotechnology. Microbiol. Mol. Biol. Rev. 66:506-577. (Abstract)
24) McLaughlin, S.G, D.G. De La Toree Ugarte, C. T. Garten, Jr., L. R. Lynd, M. A. Sanderson, V.R. Tolbert, D. D. Wolf. 2002. High-Value renewable energy from prairie grasses. Environ. Sci. Technol. 36:2122-2129. (Abstract)
23) Laser, M. D. Schulman, S. G. Allen, J. Lichwa, M. J. Antal, Jr., L R. Lynd. 2002. A comparison of liquid hot water and steam pretreatments of sugar cane bagasse for bioconversion to ethanol. Biores. Technol. 81:33-44. (Abstract)
22) Lynd, L.R., Y. Zhang. 2002. Quantitative determination of cellulase concentration as distinct from cell concentration in studies of microbial cellulose utilization: Analytical framework and methodological approach. Biotech. & Bioeng. 77:467-475. (Abstract)
21) Ozkan, M., S. G. Desai, Y. Zhang, D. M. Stevenson, J. Beane, E.A. White, M. L. Guerinot, L. R. Lynd. 2001. Characterization of thirteen newly isolated strains of anaerobic, cellulolytic, thermophilic bacteria. J. of Indust. Microbio. and Biotechnol. 27:275-280. (Abstract)
20) Allen SG, D. Schulman, J. Lichwa, M.J. Antal Jr., M. Laser, L.R. Lynd. 2001 A comparison between hot liquid water and steam fractionation of corn fiber. Ind. Eng. Chem. Res. 40:2934-2941. (Abstract)
19) Lynd, L.R., S. Baskaran, S. Casten. 2001. Salt accumulation resulting from base added for pH control, and not ethanol, limits growth of Thermoanaerobacterium thermosaccharolyticum HG-8 at elevated feed xylose concentrations in continuous culture. Biotechnol. Prog. 17:118-125. (Abstract)
18) Lynd, L.R., K. Lyford, C.R. South, G.P. Van Walsum, K. Levenson. 2001. "Evaluation of paper sludges for amenability to enzymatic hydrolysis and conversion to ethanol," TAPPI J., 84:50; full text at http://www.tappi.org.
17) Lynd, L.R., Wyman, C.E., Gerngross, T.U. Biocommodity engineering. 1999. Biotechnol. Prog. 15:777-793. (Abstract)
16) van Walsum, P., L.R. Lynd. 1998. Allocation of ATP to synthesis of cells and hydrolytic enzymes in celluloytic fermentative microorganisms: Bioenergetics, kinetics, and bioprocessing. Biotech. Bioeng. 58:316-320. (Abstract)
15) Lynd, L.R. 1997. Cellulose Ethanol: Technology in Relation to Environmental Goals and Policy Formulation. Peer-reviewed chapter, J. DeCicco and M. DeLucchi, eds. Transportation, Energy, and Environment: How Far Can Technology Take Us? American Council for an Energy-Efficient Economy Press, Washington. p 109-134. (Summary)
14) DeCicco, J., L.R. Lynd. 1997. Combining Vehicle Efficiency and Renewable Biofuels to Reduce Light Vehicle Oil Use and CO2 Emissions. Peer-reviewed chapter in, J. DeCicco and M. DeLucchi, eds. Transportation, Energy, and Environment: How Far Can Technology Take Us? American Council for an Energy-Efficient Economy Press, Washington. p. 75-108. (Summary)
13) van Walsum, G.P., S.G. Allen, M.J. Spencer, Mark S. Laser, Michael J. Antal, Lee R. Lynd. 1996. Conversion of lignocellulosics pretreated with liquid hot water to ethanol. Appl. Biochem. Biotechnol. 57/58:157-170. (Abstract)
12) Klapatch, T.R., A.L. Demain, L.R. Lynd. 1996. Restriction endonuclease activity in Clostridium thermocellum and Clostridium thermosaccharolyticum. Appl. Microbiol. Biotechnol. 45:127-131. (Abstract)
11) Lynd, L.R. 1996. Overview and evaluation of fuel ethanol from cellulosic biomass: Technology, economics, the environment, and policy. Ann. Rev. Energy Environ. 21:403-465. (Abstract)
10) Lynd, L.R., R.T. Elander, C.E. Wyman. 1996. Likely features and costs of mature biomass ethanol technology. Appl. Biochem. Biotechnol. 57/58:741-761. (Abstract)
9) Baskaran, S., Hyung-Jun Ahn, L.R. Lynd. 1995. Investigation of the ethanol tolerance of Clostridium thermosaccharolyticum in continuous culture. Biotechnol. Prog. 11:276-281. (Abstract)
8) South, C.R., D.A. Hogsett, L.R. Lynd. 1995. Modeling simultaneous saccharification and fermentation of lignocellulose to ethanol in batch and continuous reactors. Enz. Microb. Technol., 17:797-803. (Abstract)
7) Tyurin, M.V. 1992. Method for cell suspension treatment with electric current, and an apparatus for the treatment. Russian patent 2005776.
6) South, C.R., L.R. Lynd. 1994. Analysis of conversion of particulate biomass to ethanol in continuous solids retaining and cascade bioreactors. Appl. Biochem. Biotechnol. 45/46:467-481. (Abstract)
5) South, C. R., D.A. Hogsett, L.R. Lynd. 1993. Continuous fermentation of cellulosic biomass to ethanol. Appl. Biochem. Biotechnol. 39/40:587-600.
4) Lynd, L.R., J.H. Cushman, R.J. Nichols, C.E. Wyman. 1991. Fuel ethanol from cellulosic biomass. Science. 251:1318-1323. (Abstract)
3) Lynd, L.R., H.E. Grethlein, R.H. Wolkin. 1989. Fermentation of cellulosic substrates in batch and continuous culture by Clostridium thermocellum. Appl. Environ. Microbiol. 55:3131-3139. (Abstract)
2) Lynd, L.R. 1989. Production of Ethanol from Lignocellulosic Materials Using Thermophilic Bacteria: Critical Evaluation of Potential and Review. Pp 1-52 In A. Fiechter, ed., Advances in Biochemical Engineering/Biotechnology, Vol. 38, Springer-Verlag, Heidelberg. (Abstract)
1) Lynd, L. R., R.H. Wolkin, H.E. Grethlein. 1987. Continuous fermentation of pretreated hardwood and avicel by Clostridium thermocellum. Biotechnol. Bioeng. Symp. Series. 17:265- 274.