Method for glucose production with improved recovery and reuse of enzyme

• US 20040053373A1
• Filed: 09/24/2003
• Published: 03/18/2004
• Est. Priority Date: 09/25/2000
• Status: Active Grant

• Current Assignee: Iogen Energy Corporation (Iogen Corporation)
• Original Assignee: Iogen Energy Corporation (Iogen Corporation)
• Inventors: Tolan, Jeffrey S., White, Theresa C., Donaldson, Jennifer, Foody, Brian
• Granted Patent: US 7,419,809 B2
• Time in Patent Office: Days
• US Class Current: 435/105
• CPC Class Codes: C12N 9/2437 Cellulases (3.2.1.4; 3.2.1.74; 3.2.1.91; 3.2.1.150)
• CPC Class Codes: C12P 19/02 Monosaccharides 2-ketogulonic acid C12P7/60
• CPC Class Codes: C12P 19/14 produced by the action of a carbohydrase (EC 3.2.x), e.g. by alpha-amylase , e.g. by cellulase, hemicellulase
• CPC Class Codes: C12Y 302/01004 Cellulase (3.2.1.4), i.e. endo-1,4-beta-glucanase
• CPC Class Codes: C12Y 302/01091 Cellulose 1,4-beta-cellobiosidase (3.2.1.91)

Abstract

The present invention pertains to a method of converting cellulose to glucose by treating a pretreated lignocellulosic substrate with an enzyme mixture comprising cellulase enzyme and a modified CBHI, wherein the modified CBHI is present in the enzyme mixture at an amount relative to all CBHI-type enzymes from about 15% to about 100% (w/w), dpending upon the modified CBH1 used. The pretreated lignocellulosic substrate is selected from the group consisting of agricultural residues, residues after starch or sugar removal, dedicated ethanol crops, forestry products, and pulp and paper products, or combinations thereof.

Claims

1. A method of converting cellulose to glucose comprising treating a pretreated lignocellulosic substrate with an enzyme mixture comprising cellulose enzymes and a modified CBHI, wherein said cellulase enzymes are selected from the group consisting of EG, CBH and β

-glucosidase and said modified CBHI is present in said enzyme mixture at an amount relative to all CBHI-type enzymes of from about 55 to about 100 wt %.

2. The method of claim 1 wherein said amount of modified CBHI in said enzyme mixture is from about 70 to about 100 wt % relative to all CBH1-type enzymes.

3. The method of claim 1 wherein said amount of modified CBHI in said enzyme mixture is from about 80 to about 100 wt % relative to all CBH1-type enzymes.

4. The method of claim 1, wherein said modified CBHI is selected from the group consisting of CBHI core, CBHI core plus linker, CBHI with inactivated cellulose binding domain, and a combination thereof.

9. The method of claim 1, wherein said modified CBHI is recovered following said step of treating.

10. The method of claim 9, wherein said modified CBHI is reused following said step of recovery.

13. The method of claim 1, wherein said pretreated lignocellulosic substrate is selected from the group consisting of agricultural residues, residues after starch or sugar removal, dedicated ethanol crops, forestry products, and pulp and paper products or combinations thereof.

14. The method of claim 13 wherein:

said agricultural residues are selected from the group consisting of corn stover, wheat straw, barley straw, and soybean stover;

said residues after starch or sugar removal are selected from the group consisting of oat hulls, rice hulls, sugar cane bagasse, and corn fibre;

said dedicated ethanol crops are selected from the group consisting of switch grass, miscanthus, cord grass, and rye grass;

said forestry products are selected from the group consisting of hardwood, softwood, Eucalyptus, and sawdust; and

said pulp and paper products is solka floc.

17. The method of lignocellulosic hydrolysis of claim 1, wherein said lignocellulosic substrate is characterized as having an E_I of at least about 0.5, and an R_I from about 45 to about 100.

18. The method of claim 17, wherein said E_I is from about 1.0 to about 4.0, and said R_I is from about 60 to about 100.

19. The method of claim 1 wherein said pretreated lignocellulosic substrate is characterized as having an E_I of from about 0.5 to about 4.0.

20. The method of claim 19, wherein said E_I is from about 0.7 to about 2.0.

21. The method of claim 19, wherein said E_I is from about 0.8 to about 1.5.

22. The method of claim 1, wherein the amount of cellulose in the pretreated lignocellulosic substrate is present in the enzyme mixture at a concentration of about 1 wt % to about 25 wt %.

23. The method of claim 1, wherein the amount of cellulose in the pretreated lignocellulosic substrate is present in the enzyme mixture at a concentration of about 10 wt % to about 16 wt %.

30. The method of claim 1, wherein said pretreated lignocellulosic substrate is pretreated using an acid steam cook.

38. The method of claim 1 wherein said modified CBHI is a modified Trichoderma CBHI.

5. A method of converting cellulose within a cellulosic substrate into glucose comprising treating a pretreated lignocellulosic substrate with an enzyme mixture comprising modified CBHI and two or more of CBHI, CBHII, EG I, EG II, β

-glucosidase, wherein said modified CBHI is present in said enzyme mixture at an amount relative to all CBHI-type enzyme of from about 55 to about 100 wt %.

6. The method of claim 5, wherein said amount of modified CBHI in said enzyme mixture is from about 70 to about 100 wt % relative to all CBH1-type enzymes.

7. The method of claim 6, wherein said amount of modified CBHI in said enzyme mixture is from about 80 to about 100 wt % relative to all CBH1-type enzymes.

8. The method of claim 5, wherein said modified CBHI is selected from the group consisting of CBHI core, CBHI core plus linker, CBHI with inactivated cellulose binding domain, and a combination thereof.

11. The method of claim 5, wherein said modified CBHI is recovered following said step of treating.

12. The method of claim 11, wherein said modified CBHI is reused following said step of recovery.

39. The method of claim 5 wherein said modified CBHI is a modified Trichoderma CBHI.

15. A method of converting cellulose to glucose comprising treating a pretreated lignocellulosic substrate with an enzyme mixture comprising cellulase enzymes and modified CBHI for a period of time sufficient to hydrolyze said cellulose, wherein said cellulase enzymes are selected from the group consisting of EG, CBH and β

-glucosidase and said modified CBHI is present in said enzyme mixture at an amount relative to all CBHI-type enzymes of from about 55 to about 100 wt %; and

recovering said modified CBHI from said enzyme mixture.

16. The method of claim 15, wherein said modified CBHI is selected from the group consisting of CBHI core, CBHI core plus linker, CBHI with inactivated cellulose binding domain, and a combination thereof.

40. The method of claim 15 wherein said modified CBHI is a modified Trichoderma CBHI.

24. A method for hydrolyzing cellulose to glucose comprising treating a pretreated lignocellulosic substrate characterized with an E_I of from about 0.5 to about 4, with an enzyme mixture comprising cellulase enzyme with CBHI-type enzymes, wherein said CBHI-type enzymes comprise about 20% to about 100 wt % modified CBMI.

25. The method of claim 24, wherein said modified CBHI is selected from the group consisting of CBHI core, CBHI core plus linker, CBHI with inactivated cellulose binding domain, and a combination thereof.

26. The method of claim 24, wherein said step of recovery comprises using an ultrafiltration membrane.

27. The method of claim 24 wherein the amount of cellulose in the pretreated lignocellulosic substrate concentration is from about 10% to about 16 wt %.

28. The method of claim 24, wherein said pretreated lignocellulosic substrate is selected from the group consisting of agricultural residues, residues after starch or sugar removal, dedicated ethanol crops, forestry products, and pulp and paper products or combinations thereof.

29. The method of claim 28 wherein:

said agricultural residues are selected from the group consisting of corn stover, wheat straw, barley straw, soybean stover;

said residues after starch or sugar removal are selected from the group consisting of oat hulls, rice hulls, sugar cane begass, and corn fibre;

said dedicated ethanol crops are selected from the group consisting of switch grass, miscanthus, cord grass, and rye grass;

said forestry products are selected from the group consisting of hardwood, softwood, Eucalyptus, and sawdust; and

said pulp and paper products is solka floc.

41. The method of claim 24 wherein said modified CBHI is a modified Trichoderma CBHI.

31. A method of converting cellulose within a cellulosic substrate into glucose comprising;

treating a pretreated lignocellulosic substrate with an enzyme mixture comprising CBHI type enzyme, and two or more of CBHII, EG I, EG II, and β

-glucosidase, wherein CBHI core is present in said enzyme mixture at an amount relative to all CBHI-type enzyme of from about 15 to about 100 wt %.

32. The method of claim 31, wherein said CBHI core in said enzyme mixture is from about 50 to about 100 wt % relative to all CBH1-type enzymes.

33. The method of claim 31, wherein said CBHI core in said enzyme mixture is from about 80 to about 100 wt % relative to all CBH1-type enzymes.

42. The method of claim 31 wherein said CBHI core is obtained from Trichoderma CBHI.

34. A method for converting cellulose to glucose comprising treating a pretreated lignocellulosic substrate with an enzyme mixture comprising cellulase enzyme and CBHI-core plus linker enzyme, wherein said CBHI-core plus linker enzyme is present in said enzyme mixture in an amount of about 50 % to about 100 wt %, relative to all CBHI-type enzymes.

35. The method of claim 34, wherein said amount of CBHI-core plus linker in said enzyme mixture is from about 70 % to about 100 wt % relative to all CBH1-type enzymes.

36. The method of claim 35, wherein said amount of CBHI-core plus linker in said enzyme mixture is from about 80 % to about 100 wt % relative to all CBH1-type enzymes.

43. The method of claim 34 wherein said CBHI core plus linker is obtained from Trichoderma CBHI.

37. A method for converting cellulose to glucose comprising treating a pretreated lignocellulosic substrate with an enzyme mixture comprising CBHI with an inactive cellulose binding domain.