1. 1 AFEX Treatment on Poplar and Hydrolysis Balan Venkatesh, Shishir Chundawat and Bruce E. Dale BCRL, Michigan State University (www.everythingbiomass.org) 2 nd March 2006 (Integration of Leading Biomass Pretreatment Technologies, CAFI-2) Department of Chemical Engineering and Material Science

2. 2 Agenda Optimizing the AFEX pretreatment conditions (higher temperature and ammonia loading) for low lignin (small batch) and high lignin (large batch) Poplar Effect of water soaking before AFEX and water washing after AFEX Enzymatic hydrolysis using cellulases, hemicellulase and additives like surfactant

3. 3 Experimental Conditions: Water soaking: Ten times the weight of biomass (untreated) for 24 hrs AFEX condition (as given under each figure) Hydrolysis reaction condition: 50 o C, pH 4.8, 90 rpm In all experiments 15 FPU/g glucan (31.3 mg/g glucan) of Spezyme and in some reaction of multifect xylanase (3.1 mg/g glucan) was used In some reactions 0.35 g surfactant /g glucan (Tween 80) was used for 15 ml reaction volume. Hydrolyzed products were quantitatively analyzed by HPLC Calculations were done based on theoretical glucan (45.1%) and xylan (17.8%) content.

4. 4 Effect of ammonia loading on 24 h water soaked (before AFEX) High Lignin Poplar C- Cellulase X- Xylanase A- Additive UT- Untreated

5. 5 1:1 (BM:Ammonia)1:1.5 (BM:Ammonia) Effect of Temperature and ammonia loading on Low Lignin Poplar C- Cellulase X- Xylanase A- Additive UT- Untreated BM- Biomass

6. 6 Effect of Temperature and ammonia loading on 24 h water soaked (before AFEX) Low Lignin Poplar ?? C- Cellulase X- Xylanase A- Additive UT- Untreated BM- Biomass

7. 7 For Corn Stover (UT) 6%, (AT) 13% (weight loss based on DBM) For Low Lignin Poplar (UT) 3%, (AT) 7% (weight loss based on DBM) Mass Balance For Water Washing UT- Untreated AT- AFEX treated CS- Corn stover

8. 8 UT- Untreated AT- AFEX treated UG- Ungrounded GR- Ground C- Cellulase X- Xylanase A- Additive (Tween 80) BM- Biomass Effect of Grinding on Enzymatic hydrolysis

9. 9 Future work: Modify the pretreatment condition for higher glucan conversions Effect of various additives and enzyme combinations on hydrolysis Wash stream analysis for lignin monomers and xylo-oligosaccharides by LC-MS, GC-MS Inhibitory effect of wash stream components on enzymes Present results: Increasing pretreatment temperature, improves glucan conversion Water soaking before AFEX further enhances glucan conversion Grinding aids AFEX pretreatment effectiveness and improves enzymatic hydrolysis as well