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2004 R&D 100 Award Winner

Enzymatic Hydrolysis of Biomass Cellulose to Sugars

Principal Developers: Dr. Michael Himmel, Dr. James McMillan, and Dr. Rafael Nieves, National Renewable Energy Laboratory; Dr. Colin Mitchinson and team, Genencor International; Dr. Joel Cherry and team, Novozymes Biotech, Inc.

Inexpensive sugars from biomass for making fuels and chemicals. NREL and its partners — Genencor International and Novozymes Biotech, Inc. — have developed an enzymatic hydrolysis technology that inexpensively breaks down the cellulose fraction of lignocellulosic biomass to sugars. The sugars can then serve as platform chemicals from which to derive fuels, chemicals, plastics, fibers, detergents, pharmaceuticals, and many other products — all sustainably. Such a concept, known as biorefining, is analogous to petroleum refining except it is based on renewable feedstocks and could help America offset its use of petroleum-derived fuels and chemicals.

Before the advent of our technology, the process for breaking down cellulose to sugars was very expensive — too expensive to compete with the technology used to break down the starch in corn kernels to sugars or to compete with breaking down hydrocarbons in petroleum for the production of the many fuels, chemicals, and products that help run the modern economy.

Lignocellulosic biomass consists of cellulose, hemicellulose, and lignin, with cellulose constituting up to 50% of the total mass. Like starch and sugar, hemicellulose and cellulose are carbohydrates (compounds of carbon, hydrogen, and oxygen). The sugars of which they are made are linked together in long chains called polysaccharides, which form the structural portion of plant cell walls. Unraveling these complex polymeric structures is the key to economic biorefining.

Cellulose microfibrils consist of a crystalline structure of thousands of strands, each of which contains hundreds of glucose sugar molecules. These microfibrils are wrapped in a sheath of hemicellulose and lignin, which protects the cellulose from microbial attack. Hemicellulose is relatively easy to break down into its component sugars using heat, pressure, and dilute acid. This "dilute acid hydrolysis" pretreatment step also disrupts the hemicellulose/lignin sheath around the cellulose, making the cellulose accessible to further hydrolysis.

To hydrolyze the cellulose, NREL and its partners developed a technology that employs a cocktail of three types of cellulase enzymes — endoglucanase, exoglucanase, and betaglucosidase. First, an endoglucanase attacks one of the cellulose chains within the crystal structure, breaking the strand via hydrolysis, and thereby exposing two new chain ends. During this hydrolysis, a molecule of water is consumed and one of the chain ends chemistry becomes "reducing" and the other "non-reducing." Then an exoglucanase attaches to a loose chain end, physically pulls the cellulose chain away from the crystal structure, and then proceeds to work its way down the chain, breaking off cellobiose (a dimeric sugar comprised of two glucose molecules) as it goes. (Actually, there are two types of exoglucanase — a cellobiohydrolase I (CBH I) attaches to the "reducing" end and a cellobiohydrolase II (CBH II) attaches to the "non-reducing" end.) Finally, a betaglucosidase splits the cellobiose molecule into two separate glucose molecules, making them available for processing into chemicals or fuels.

Until the advent of our breakthrough technology, other methods of hydrolyzing cellulose to sugars — including strong acid hydrolysis and older versions of enzymatic hydrolysis — were inefficient, expensive, and had low sugar yields. But, by improving the pretreatment process, engineering new enzymes that are exceptional at breaking down cellulose, and optimizing enzyme production, NREL and its partners developed a technology that is efficient, has high sugar yields, and that has already dropped the cost of cellulose hydrolysis by 20-fold. Moreover, this technology holds the promise of decreasing the cost by another order of magnitude or more.