Product Information DESCRIPTION ACCELLERASE® BG beta-glucosidase is designed as an accessory product to supplement whole cellulases deficient in beta-glucosidase and work synergistically to achieve high glucan conversion for the lignocellulosic biomass processing industry. ACCELLERASE® BG enzyme can effectively convert cellobiose into fermentable mono-saccharides like glucose. Cellobiose is also an inhibitor of cellulase enzymes, so low levels are desirable. Supple-mentation with ACCELLERASE® BG enzyme can result in faster and higher ethanol production. ACCELLERASE® BG is a beta-glucosidase produced from a genetically modified strain of Trichoderma reesei. The production host is removed at the end of the controlled fermentation and the product is concentrated and formu-lated.

TYPICAL CHARACTERISTICS Activity: 3000 pNPG U/g (minimum) Appearance: Clear amber liquid pH: 5.0 - 6.0 The activity of ACCELLERASE® BG enzyme is expressed in pNPG units. One pNPG unit denotes 1 μmol of Nitrophe-nol liberated from para-nitrophenyl-B-D-glucopyranoside per minute at 50°C (122°F) and pH 4.8. Detailed assay method is available upon request.

APPLICATION RECOMMENDATIONS ACCELLERASE® BG accessory enzyme, when added to other cellulase products deficient in beta-glucosidase, will enhance saccharification of glucan in lignocellulosic bio-mass and increase the yield of fermentable glucose by alleviating product inhibition to the cellulolytic enzymes. ACCELLERASE® BG enzyme can be used with whole cellu-lases in hydrolysis of various feedstocks, including paper pulp, corn stover and cob, sugarcane bagasse, wheat straw, wood chips, waste paper and many others obtained from different pretreatment technologies such as dilute acid, ammonia fiber-expansion (AFEX) and steam expan-sion. ACCELLERASE® BG enzyme can be used in simulta-neous saccharification and fermentation (SSF), two step sequential hydrolysis and fermentation (SHF), or hybrid saccharification and fermentation (HSF) configurations. Please be aware that the pH and temperature stability optima and limits of the enzyme in use will depend upon the other operating parameters and your process configuration. Saccharification performance may be enhanced by the addition of other Genencor® enzymes depending on the composition of the pretreated feed-stock.

Accessory Beta-Glucosidase for Biomass Hydrolysis

ACCELLERASE® BG

DOSAGE GUIDELINES ACCELLERASE® BG enzyme is generally added to other cellulase products to improve hydrolysis performance. The optimum dosage levels of ACCELLERASE® BG enzyme will vary considerably with different substrates and their associated pretreatment technologies and con-ditions, and other enzyme additions, if applicable. Oper-ating conditions such as pH, temperature and reaction time may also affect enzyme performance. ACCELLE-RASE® BG enzyme dosage rate of 0.009 to 0.18 mL per gram cellulose or roughly 0.0045 to 0.09 mL per gram of biomass (depending on biomass composition), in addi-tion to, or in partial substitution of, total enzyme, is rec-ommended as a starting point for enzyme blending opti-mization. Small-scale experiments are recommended to determine optimum enzyme dosage and blends in each system. See Figures 1-3 for examples of such experi-ments.

Figure 1: Cellobiose concentration profiles for a washed acid-pretreated sugarcane bagasse (APB) at 7% cellulase loading using SPEZYME® CP whole cellulase and ACCELLERASE® BG enzyme at 50°C and pH 5.0. Experiments were performed in 100g batch-scale experiments in 500ml shake-flasks. Final total protein loading is the same in every case.

Together we can fuel the future with biomass.

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PACKAGING ACCELLERASE® BG enzyme is available in various package sizes. Please consult your Genencor® representative for detailed informa-tion.

STORAGE It is advisable to store ACCELLERASE® BG enzyme under refriger-ated conditions of 4 - 10°C (39 - 50°F), sheltered against direct sunlight. Storage above 20°C (70°F) should be avoided.

SAFETY & ENZYME HANDLING Inhalation of enzyme dust and mists should be avoided. In case of contact with the skin or eyes, promptly rinse with water for at least 15 minutes. For detailed handling information, please refer to the appropriate Material Safety Data Sheet, the Enzyme Technical Association (ETA) handbook Working Safely With Enzymes, and the Association of Manufacturers and Formulators of Enzyme Products (Amfep) handbook Guide to the Safe Handling of Microbial Enzyme Preparations. All are available from Genencor®.

TECHNICAL SERVICE Information covering specific applications of this product is available. Genencor® will work with customers to enhance processes and solve problems. Let us know what you need and we will assist you.

Figure 2: Glucose concentration profile for a washed acid-pretreated sugarcane bagasse (APB) and a pretreated softwood pulp (PSP)* at 7% cellulose loading using SPEZYME® CP enzyme and ACCELLERASE® BG enzyme at 50°C and pH 5.0. Saccharifica-tion using ACCELLERASE® 1500 whole cellulase (0.36 mL product/g cellulose) is included for comparison. Experiments were performed in 100 g batch-scale experiments in 500 mL shake-flasks. Final total protein loading is the same in every case. When adding ACCELLERASE® BG enzyme to whole cellu-lases, such as SPEZYME® CP, overall glucan conversion can be greatly improved and is comparable to the performance of ACCELLERASE® 1500 enzyme.

Figure 3: Simultaneous saccharification and fermentation (SSF) of a pretreated softwood pulp* at 7% cellulose loading, pH 5.0 and 38°C with THERMOSACC® DRY yeast (Milwaukee, WI) using either SPEZYME® CP (0.21 mL product/g cellulose), ACCELLERASE® 1500 (0.36 mL product/g cellulose), or a blend of SPEZYME® CP enzyme (0.18 mL product/g cellulose) and ACCELLERASE® BG enzyme (0.05 mL product/g cellulose). 100 g batch-scale experiments in 500 mL shake-flasks. Final total protein loading is the same in every case. * Data on pretreated softwood pulp was generated in research funded by Agence Nationale de la Recherche, France (ANR-05-BIOE-007) through L'Agence de l"Environnement et de la Maîtrise de l'Energie (ADEME 0501 C0099).

EFFECT OF pH AND TEMPERATURE ACCELLERASE® BG enzyme has the best operational stability in the following ranges: Temperature: 30 - 55°C (122 - 167°F) pH: 4.0 – 6.0 Whereas long operational times are possible at the low end of this temperature range, the higher temperatures will limit the effective periods of operation. ACCELLERASE® BG enzyme is easily inactivated at tempera-tures above 70°C (158°F) or at pH levels above 7.0 or below 4.0.

REV0409 3048 ACCELLERASE BG 01

Contact information: USA and Canada Rochester, New York Telephone: 1-800-847-5311 Telefax: +1-585-244-4544 Europe, Africa and Middle East Leiden, The Netherlands Telephone: +31-71-5686-168 Telefax: +31-71-5686-169 Latin America Buenos Aires, Argentina Telephone: +54-11-5199-9550 Telefax: +54-11-5199-9559

© Danisco US Inc. 2009. GENENCOR and ACCELLERASE are registered trademarks of Danisco US Inc. or its affiliated companies. USE AND/OR SALE OF THIS PRODUCT MAY BE COVERED BY ONE OR MORE OF THE PATENTS: U.S. 6,022,725 AND FOREIGN EQUIVALENTS, EP 056 2003 B1, CANADIAN PATENT 2,097,180. PATENTS PENDING IN THE U.S. AND OTHER COUNTRIES.

The information contained in this product literature is, to the best of our knowledge, true and accurate and the product as sold is in conformance with the specifications set forth herein as determined by the assay methods described. Due to conditions of use, technical errors or omissions, improper handling or storage beyond our control, Danisco US Inc. hereby DISCLAIMS ANY EXPRESS OR IMPLIED WARRANTIES, INCLUDING THE IMPLIED WARRANTIES OF MERCHANTABILITY OR FITNESS FOR A PARTICULAR PURPOSE. Danisco US Inc. shall not be liable for any incidental, consequential, or special damages resulting in any way from the furnishing, performance, or use of this product literature or the product described herein.

Nothing contained herein shall be construed as a representation by Danisco US Inc. that the use or resale of the product or processes described herein will not violate any rules or regulations or infringe upon patents or other intellectual property rights of third parties or that the recommendations and usage suggestions described constitute any authorization or inducement to infringe any such rights.

Asia/Pacific Singapore Telephone: +65-6511-5600 Telefax: +65-6511-5666 Shanghai, P.R. China Telephone: +86-21-2307-9588 Telefax: +86-21-2307-9599 Mumbai, India Telephone: +91-22-2825-8713 Telefax: +91-22-2825-8733 Web Address www.genencor.com

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