1. Investigating and Modeling Natural Biodegradation System in Soil Mythreyi Chandoor, Deepak Singh and Shulin Chen Bioprocessing and Bioproduct Engineering Laboratory, Department of Biological Systems Engineering Washington State University.

2. Outline Aim and importance of the project Background – Hypothesis of the project Experimental:  Microbiology  Chemical analysis of lignocellulose degradation in soil. Modeling  Lignocellulose degradation in soil  Application Acknowledgements

3. Aim and importance of the project Demand for an Alternative Fuel The U.S. ethanol consumption is forecast to increase from 5.6 billion gallons last year to 13.5 billion gallons in 2012, (Thomson Reuters, 2009). What are the challenges ? One of the challenges lies in the deconstruction of lignin part of the biomass to release sugars. Need for novel pretreatment technology !

4. Delignification, repolymerization Humus formation Proteins Hypothesis: Lignocellulosic degradation system in soil Parallel degradation of cellulose and hemicellulose

5. Lignin –Major Linkages in the structure Lignin surrounds cellulose microfilbrils,there by providing strength to the cell wall. Highly heterogeneous material,requires Oxygen for its degradation /modification.

6. To understand the biodegradation of lignocellulose in soil To model the biodegradation of lignocellulose in soil Design the pretreatment system Aim of the Project

7. Methodology

8. Scanning Electron Microscopy (SEM)

10. C2,C3,C5 of cellulose C4 of amorphous and crystalline cellulose 4 weeks 8 weeks 12 weeks 16 weeks Solid State NMR Analysis Syringol Guiacyl Sinapyl, coniferyl

11. Batch samples for every four weeks % Concentration of the total compound Pyrolysis-GC/MS Analysis

12. Thermogravitometric Analysis Cellulose Lignin Hemicellulose 40 60 80 100 20 Weight percent (%)

13. [S]+[X]+O 2 + H 2 O soil pH [p] +[s 1 ] + co 2 +[X] Input = output +accumulation – generation General Equation for the Soil Degradation system

14. d(S 1 ) / d(t) = -Vb 1 *S 1 *X1/(Ks 1 +S 1 ) S 1 (0) = 0.71 # weight in gm/gm #Cellulose Balance d(S 2 ) / d(t) = -Vb 2 *S 2 *X 2 /(Ks 2 +S 2 ) S 2 (0) = 0.48 # weight in gm/gm #Hemicellulose Balance d(S 3 ) / d(t) = -Vb 3 *S 3 *X 3 /(Ks 3 +S 3 ) S 3 (0) = 0.28 # weight in gm/gm #Lignin Balance

15. #Microbial Growth corresponding to Cellulose d(X 1 ) / d(t) = μ 1 *X 1 X 1 (0) = 0.1 #Microbial Growth corresponding to Hemicellulose d(X 2 ) / d(t) = μ 2 *X 2 X 2 (0) = 0.1 #Microbial Growth corresponding to Lignin d(X 3 ) / d(t) = μ 3 *X 3 X 3 (0) = 0.1 µ 1 =µ max1 *S 1 /(Ks 1 +S 1 )-∆ 1 µ 2 =µ max2 *S 2 /(Ks 2 +S 2 )-∆ 2 µ 3 =µ max3 *S 3 /(KS 3 +S 3 )-∆ 3

16. t(0) = 0 t(f) = 3360 Considering the values as follows ; µmax1=0.08 µmax2=0.05 μmax3=0.03 system. ∆ 1 =0.001 ∆ 2 =0.001 ∆ 3 =0.001 We derived an relation using polymath which defines the degradation pattern in the soil Initial and final time periods of Incubation

17. Time (in hours ) Initial Substrate concentration in gm / gm We derived an relation using polymath which defines the degradation pattern in the soil system. Modeling

18. Application of the Model The determination of the exact relation between the various factors with respect to the specific incubation period would be helpful in developing a model which would predict the specific ratio of cellulose, hemicellulose and lignin apart from other factors involved such as pH, temperature and other organic compounds. Thus providing a suitable mechanism for the pretreatment technology !

19. Conclusion Scientific Contribution : Specific conditions which relates to the changes in the relative composition of cellulose,hemicellulose and lignin, Chemical changes in the lignin structure with respect to time as a result of which lignin modification in the soil takes place during the natural degradation process was determined Implication: As the pathway of lignin degradation in soil is quite different this would provide us a scope for a novel method for lignin degradation process. Further work would involve understanding different factors which play an important role during the lignin modification and degradation.

20. I would like to thank Dr. Ann Kennedy USDA-ARS Soil Scientist/Adj. Prof. Crop and Soil Sciences,WSU. Dr. Greg Helms, NMR Center Director,WSU. Dr. Manuel Garcia-Perez. Assistant Professor / Scientist. Biological Systems Engineering,WSU. Dr. Bill, Assistant manager,NMR Center,WSU. And my Advisor … Dr. Shulin Chen, Professor/Scientist. Department of Biological Systems Engineering,WSU.

21. My Team …

22. Thank you … Any Questions ?