Production of chemicals from lignocellulose, biomass or sugars

• US 20050250192A1
• Filed: 03/04/2004
• Published: 11/10/2005
• Est. Priority Date: 03/04/2004
• Status: Active Grant

• Current Assignee: University of Florida Research Foundation, Incorporated (State University System of Florida)
• Original Assignee: University of Florida Research Foundation, Incorporated (State University System of Florida)
• Inventors: Shanmugam, Keelnatham T., Ingram, Lonnie O'Neal, Patel, Milind A., Ou, Mark S., Harbrucker, Roberta
• Granted Patent: US 7,098,009 B2
• Time in Patent Office: Days
• US Class Current: 435/139
• CPC Class Codes: C12N 1/205 Bacterial isolates
• CPC Class Codes: C12N 15/52 Genes encoding for enzymes or proenzymes
• CPC Class Codes: C12N 9/1029 transferring groups other than amino-acyl groups (2.3.1)
• CPC Class Codes: C12P 7/10 substrate containing cellulosic material
• CPC Class Codes: C12P 7/18 polyhydric
• CPC Class Codes: C12P 7/46 Dicarboxylic acids having four or less carbon atoms, e.g. fumaric acid, maleic acid
• CPC Class Codes: C12P 7/56 Lactic acid
• CPC Class Codes: C12R 2001/07 Bacillus
• CPC Class Codes: Y02E 50/10 Biofuels, e.g. bio-diesel

Abstract

The subject invention relates to newly isolated organisms from nature that produce L(+)-lactic acid high yield from hexose and pentose sugars found in biomass. Organisms and processes or methods for the production of lactic acid and other industrially important chemicals from cellulose and hemicellulose are also provided.

Claims

1. An isolated Gram positive organism that produces L(+) lactic acid at high yield from hexose or pentose sugars or combinations of hexose and pentose sugars, wherein said organism has not been genetically modified or transformed, has been isolated from nature, is phylogenetically distinct from Bacillus coagulans and said organism is not Bacillus smithii of Bacillus coagulans, strain IDSp.

2. The organism according to claim 1, wherein said organism comprises at least one of the following characteristics:

a) able to grow under anaerobic conditions;

b) able to grow and ferment glucose, xylose, arabinose, galactose, mannose, sucrose, cellobiose, or various combinations of these carbohydrate sources;

c) able to grow in minimal-salts medium with and without corn steep liquor;

d) able to grow in, and ferment, sugar cane bagasse hemicellulose hydrolysate, crystalline cellulose;

e) performance of simultaneous saccharification and fermentation (SSF) of crystalline cellulose;

f) classification into a unique phylogenetic group of organisms subject on the basis of 16S rRNA sequences and wherein said organisms has a similarity index of at least 0.97 as compared to at least one, any combination, or all, of the sequences of SEQ ID NOs;

3-42;

g) generation of L(+)-lactic acid yields in excess of 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, or 99%, wherein the L(+)-lactic acid produced by the organisms has an optical purity of at least 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, or 99%;

h) spore-forming organism;

i) classification as a member of the family Bacillacene;

or

j) classification as a member of the genus Bacillus.

11. A method of producing an industrially useful chemical comprising the steps of:

1) providing

a non-recombinant organism according to claim 1;

and

2) culturing said microorganism in the presence of at least one carbon source capable of being converted to said industrially useful chemical under conditions suitable for the production of said industrially useful chemical.

12. The method according to claim 11, further comprising the step of recovering the industrially useful chemical.

13. The method according to claim 11, wherein said industrially useful chemical is L(+)-lactic acid.

14. The method, according to claim 11, wherein said carbon source is a hexose or pentose sugar.

20. The method according to claim 11, wherein both a recombinant or transformed organism and a non-recombinant organism are provided for the production of said industrially useful chemical.

22. The organism according to claim 1, wherein said organism utilizes the pentose phosphate pathway for xylose fermentation.

23. The organism according to claim 2, wherein said organism is classified into a unique phylogenetic group of organisms on the basis of 16S rRNA sequences and wherein said organism has a similarity index of at least 0.97 as compared to at least one, any combination, or all, of the sequences of SEQ ID NOs:

3-42.

24. The organism according to claim 22, wherein said organism is classified into a unique phylogenetic group of organisms on the basis of 16S rRNA sequences and wherein said organism has a similarity index of at least 0.97 as compared to at least one, any combination, or all, of the sequences of SEQ ID NOs:

3-42.

3. A method of isolating an organism comprising the steps of:

a) obtaining an environmental sample from soils, compost, wood chips, food products, mulch, animal waste, or any other environmental source;

b) pre-incubating and resuspending the sample;

c) enriching for cellulose positive organisms;

d) isolating hemicellulose fermenting organisms;

e) enriching for hemicellulose fermenting organisms;

f) isolating organisms enriched in step e); and

g) analyzing said isolated organisms for desired properties, wherein said desired properties include the ability to produce L(+) lactic acid at high yield from hexose and pentose sugars and at least one property selected from the group consisting of;

a) the ability to grow under anaerobic conditions;

b) the ability to grow and ferment glucose, xylose, arabinose, galactose, mannose, sucrose, cellobiose, or various combinations of these carbohydrate sources;

c) the ability to grow in minimal-salts medium with and without corn steep liquor;

d) the ability to grow in, and ferment, sugar cane bagasse hemicellulose hydrolysate, crystalline cellulose;

e) perform simultaneous saccharification and fermentation (SSF) of crystalline cellulose;

f) classification into a unique phylogenetic group of organisms on the basis of 16S rRNA sequences;

g) generation of L(+)-lactic acid yields in excess of 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, or 99%, wherein the L(+)-lactic acid produced by the organisms has an optical purity of at least 80, 81, 82, 83. 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, or 99%;

h) spore formation;

i) classification as a member of the family Bacillaceae; and

j) classification as a member of the genus Bacillus;

or any combination of said properties.

4. An organism isolated according to the method of claim 3, provided that said organism is not B. coagulans, B. coagulans strain IDSp, or B. smithii,

25. The organism according to claim 4, wherein said organism produces L(+) lactic acid at high yield from hexose and pentose sugars and has at least one property selected from the group consisting of:

a) the ability to grow under anaerobic conditions;

b) the ability to grow and ferment glucose, xylose, arabinose, galactose, mannose, sucrose, cellobiose, or various combinations of these carbohydrate sources;

c) the ability to grow in minimal-salts medium with and without corn steep liquor;

d) the ability to grow in, and ferment, sugar cane bagasse hemicellulose hydrolysate, crystalline cellulose;

e) perform simultaneous saccharification and fermentation (SSF) of crystalline cellulose;

f) classification into a unique phylogenetic group of organisms on the basis of 16S rRNA sequences;

g) generation of L(+)-lactic acid yields in excess of 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, or 99%, wherein the L(+)-lactic acid produced by the organisms has an optical purity of at least 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, or 99%;

h) spore formation;

i) classification as a member of the family Bacillaceae; and

j) classification as a member of the genus Bacillus.

26. The organism accord claim 25, wherein said organism has the ability to ferment xylose.

27. The organism according to claim 25, wherein said organism ferments pentose via the pentose phosphate pathway.

5-10. -10. (canceled)

15-19. -19. (canceled)

21. An isolated microorganism selected from the group consisting of:

a) Bacillus isolates 17C5 (ATCC PTA-5826), 36D1 (ATCC PTA-5827), and P4-102B (ATCC PTA-5828);

b) progeny of Bacillus isolates 17C5, 36D1, and P4-102B;

c) subclones of Bacillus isolates 17C5, 36D1, and P4-102B; and

d) subcultures of Bacillus isolates 17C5, 36D1, and P4-102B.