Filamentous fungal biomats, methods of their production and methods of their use

• US 10,590,379 B2
• Filed: 06/14/2019
• Issued: 03/17/2020
• Est. Priority Date: 03/01/2016
• Status: Active Grant

• Current Assignee: Fynder Group Incorporated
• Original Assignee: Sustainable Bioproducts, Inc.
• Inventors: Kozubal, Mark A., Macur, Richard E., Avniel, Yuval C.
• Primary Examiner(s): Yamasaki, Robert J
• Application Number:
  US16/442,151
• Publication Number:
  US 20190316078A1
• Time in Patent Office: 277 Days
• Field of Search: None
• CPC Class Codes: C02F 3/347 Use of yeasts or fungi C02F3/322 takes precedence
• CPC Class Codes: C12N 1/02 Separating microorganisms from their culture media
• CPC Class Codes: C12N 1/14 Fungi culture of mushrooms A01G18/00; as new plants A01H15/00; Culture media therefor
• CPC Class Codes: C12N 1/145 Fungal isolates
• CPC Class Codes: C12R 2001/645 Fungi ; Processes using fungi
• CPC Class Codes: C12R 2001/66 Aspergillus
• CPC Class Codes: C12R 2001/77 Fusarium
• CPC Class Codes: C12R 2001/785 Mucor
• CPC Class Codes: C12R 2001/80 Penicillium
• CPC Class Codes: C12R 2001/84 Pichia
• CPC Class Codes: C12R 2001/845 Rhizopus
• CPC Class Codes: C12R 2001/885 Trichoderma

Abstract

A novel method of growing fungi is disclosed which uses an engineered artificial media and produces high density filamentous fungi biomats that can be harvested with a minimum of processing and from which fungal products such as antibiotics, proteins, and lipids can be isolated, the method resulting in lowered fungus cultivation costs for energy usage, oxygenation, water usage and waste stream production.

Claims

1. A method of producing a biomat by a surface fermentation method comprising

(a) inoculating an artificial liquid growth medium with an effective amount of a biologically pure culture of filamentous fungal strain MK7 (ATCC Accession Deposit No. PTA-10698);

(b) incubating the inoculated filamentous fungi to produce a filamentous fungal biomat on the surface of the liquid medium; and

(c) harvesting the filamentous fungal biomat,

wherein the biomat comprises at least two structurally different cell layers in contact with each other, including an upper aerial hyphae layer in contact with the atmosphere during the surface fermentation and a lower layer in contact with the liquid media during the surface fermentation, wherein the lower layer has a higher fungal density than the aerial hyphae layer.

2. The method of claim 1, wherein the artificial liquid growth media comprises a carbon source selected from the group consisting of starch, acid whey, corn steep liquor, potato waste, sugar beet waste, glycerol, dextrose, glucose, maltose, galactose, mannose, corn syrup, vegetable scraps, and cattle feedstock water.

3. The method according to claim 2, wherein the carbon source is acid whey.

4. The method according to claim 2, wherein the carbon source is glycerol.

5. The method according to claim 1, wherein the artificial liquid growth media comprises a carbon source comprising a waste selected from the group consisting of an agricultural waste, a clinical organic waste, a municipal organic waste, a food processing waste, a biofuel production waste, an industrial waste, a lignocellulosic containing waste, dairy waste, slaughterhouse waste and a combination thereof.

6. The method according to claim 1, wherein the artificial liquid growth media comprises a carbon source selected from the group consisting of a monosaccharide, a disaccharide and a combination thereof.

7. The method according to claim 1, wherein the artificial liquid growth media comprises a carbon source selected from the group consisting of glycerol, a sugar, dried wheat distillers solubles, corn steep liquor, cattle feedlot lagoon water, acid whey, vegetable wastes, corn syrup and a combination thereof.

8. The method according to claim 1, wherein the artificial growth media has an osmotic pressure of about 18.6 atm.

9. The method according to claim 1, wherein the artificial growth media has an ionic strength of about 0.368 M.

10. The method according to claim 1, wherein the artificial growth media comprises ammonium nitrate, urea, potassium dihydrogen phosphate, calcium chloride, magnesium sulfate, and yeast extract.

11. The method according to claim 10, wherein the artificial growth media further comprises ferric sulfate, zinc sulfate, manganese chloride, cobalt chloride, copper sulfate, and boric acid.

12. The method according to claim 1, wherein step (a) comprises inoculating an effective amount of planktonic cells of the filamentous fungus MK7.

13. The method according to claim 12, wherein 7.5% (volume:

volume) of planktonic cells are inoculated into the artificial growth media.

14. The method according to claim 1, wherein the biomat comprises three structurally different cell layers.

15. The method according to claim 1, wherein the biomat produced in step (b) comprises at least 25 g dry wt filamentous fungi/1 media.

16. The method according to claim 1, wherein the biomat produced in step (b) comprises at least 50 g dry wt filamentous fungi/1 media.

17. The method according to claim 1, wherein the biomat produced in step (b) comprises at least 120 g dry wt filamentous fungi/1 media.

18. The method according to claim 1, wherein the biomat produced in step (b) comprises at least 180 g dry wt filamentous fungi/1 media.

19. The method of claim 1, wherein the artificial growth media comprises a nitrogen source.

20. The method of claim 19, wherein the nitrogen source is selected from the group consisting of nitrate salts, ammonium salts, proteins, peptides, urea, waste streams comprising nitrogen, and combinations thereof.

21. The method of claim 1, wherein the artificial liquid growth media comprises carbon and nitrogen, and the ratio of C:

N is about 7.5;

1 or less.

22. The method of claim 1, wherein the artificial liquid growth media comprises carbon and nitrogen, and the ratio of C:

N is greater than about 7.5;

1.

23. The method of claim 1, wherein the biomat has a protein content of at least 40%.

24. The method of claim 1, wherein the biomat has a lipid content of at least 39%.

25. The method of claim 1, wherein the biomat comprises amino acids selected from selected from tryptophan, cysteine, methionine, alanine, arginine, aspartic acid, glutamic acid, glycine, histidine, isoleucine, leucine, phenylalanine, proline, serine, threonine, lysine, tyrosine and valine.