# US 11,111,510 B2 · Fermentation process for the production of lipids

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- Number: US 11,111,510 B2
- Document ID: US11111510B2
- Status: Granted
- Kind: B2
- Application number: 16/723,498
- Filed: 2019-12-20
- Published: 2020-04-23
- Granted: 2021-09-07
- Earliest claimed: 2014-10-31
- Assignees: Lanzatech New Zealand Limited; DBT-IOC Centre for Advanced Bio-Energy Research
- Applicants: LanzaTech New Zealand Limited; DBT-IOC Centre for Advanced Bio-Energy Research
- Inventors: Sean Dennis Simpson; Sebastian Michal Bernasek; Deepak K Tuli
- Web page: https://patentgrep.com/patent/US11111510B2

## CPC

- C12P 7/6463: obtained from glyceride producing microorganisms, e.g. single cell oil
- C12P 7/6427: Polyunsaturated fatty acids [PUFA], i.e. having two or more double bonds in their backbone
- C12P 7/6432: Eicosapentaenoic acids [EPA]
- C12P 7/6434: Docosahexenoic acids [DHA]
- C12P 7/6472: containing polyunsaturated fatty acid [PUFA] residues, i.e. having two or more double bonds in their backbone
- C12P 39/00: Processes involving microorganisms of different genera in the same process, simultaneously
- Y02E 50/10: Biofuels, e.g. bio-diesel
- C12P: FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE {(brewing of beer C12C; producing vinegar C12J; producing specific peptides or proteins C07K; producing enzymes C12N9/00; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification C12N15/00; measuring or testing processes involving enzymes or microorganisms C12Q; measuring or testing processes involving nucleic acid amplification reactions C12Q1/6844; fermentation processes to form a food composition, A21 or A23; compounds in general, see the relevant compound class, e.g. C01, C07)}
- Y02E: REDUCTION OF GREENHOUSE GAS [GHG] EMISSIONS, RELATED TO ENERGY GENERATION, TRANSMISSION OR DISTRIBUTION

## Abstract

The invention provides methods and systems for the production of lipid products from a gaseous substrate using a two stage fermentation process. The method comprises providing a gaseous substrate comprising CO, CO2 or H2 or mixtures thereof, to a first bioreactor containing a culture or one or more microorganisms, and fermenting the substrate to produce acetate. The acetate from the first bioreactor is then provided to a second bioreactor, where it is used as a substrate for fermentation to lipids by one or more microalgae.

## Claims

### Claim 1

A method for producing at least one lipid product from a gaseous substrate, the method comprising:
i. receiving a gaseous substrate comprising CO2 and H2 in a first bioreactor containing a culture of at least one microorganism in a liquid nutrient medium, and fermenting the gaseous substrate under anaerobic conditions to produce acetate;
ii. passing at least a portion of the acetate to a second bioreactor containing a culture of at least one microalgae in a liquid nutrient medium, and fermenting the acetate under aerobic conditions to produce at least one lipid product, wherein the microalgae is selected from the group consisting of Thraustochytrium, Japonochytrium, Aplanochytrium, Elina, and Labyrinthula;
iii obtaining a fermentation broth from the second bioreactor; and
iv. passing at least a portion of the fermentation broth from the second bioreactor to a separator to produce a product stream comprising at least one lipid and biomass and an acetate depleted permeate stream;
v. passing the permeate stream to an oxygen removal unit to remove oxygen and then recycling at least a portion of the resulting oxygen depleted stream to the first bioreactor.

### Claim 2 (depends on 1)

The method of claim 1, wherein the production rate of acetate in the first bioreactor is at least 10 g/L/day.

### Claim 3 (depends on 1)

The method of claim 1, wherein the at least one microorganism in the first bioreactor is selected from the group consisting of Acetobacterium, Moorella, Clostridium, Pyrococcus, Eubacterium, Desulfobacterium, Carboxydothermus Acetogenium, Acetoanaerobium, Butyribacterium, Peptostreptococcus, Ruminococcus, Oxobacter, and Methanosarcina.

### Claim 4 (depends on 1)

The method of claim 1, wherein the at least one microalgae in the second bioreactor is Thraustochytrium.

### Claim 5 (depends on 1)

The method of claim 1, further comprising converting the at least one lipid product produced in stage (ii) to a product selected from a hydrogenation-derived renewable diesel (HDRD), fatty acid methyl esters (FAME), fatty acid ethyl esters (FAEE), or biodiesel.

### Claim 6 (depends on 1)

The method of claim 1, wherein lipid production is increased in the second bioreactor by limiting at least one nutrient in the liquid nutrient medium.

### Claim 7 (depends on 6)

The method of claim 6, wherein the limited nutrient is nitrogen.

### Claim 8 (depends on 1)

The method of claim 1, wherein the lipid product is a polyunsaturated fatty acid.

### Claim 9 (depends on 8)

The method of claim 8, wherein the polyunsaturated fatty acid is an omega-3 fatty acid.

### Claim 10 (depends on 9)

The method of claim 9, wherein the omega-3 fatty acid is one or more of alpha-linolenic acid (ALA), eicosapentaenoic acid (EPA), and docosahexaenoic acid (DHA).

### Claim 11 (depends on 1)

The method of claim 1, wherein the at least one microorganism in the first bioreactor is an acetogenic bacterium.

### Claim 12 (depends on 1)

The method of claim 1, wherein the microalgae is selected from the group consisting of Japonochytrium, Aplanochytrium, Elina, and Labyrinthula.

### Claim 13 (depends on 1)

The method of claim 1, wherein the liquid nutrient medium of the second bioreactor comprises nitrogen.

### Claim 14 (depends on 13)

The method of claim 13, wherein the liquid nutrient medium of the second bioreactor comprises an acetic acid to nitrogen ratio of at least 10:1.

### Claim 15 (depends on 13)

The method of claim 13, wherein the liquid nutrient medium of the second bioreactor comprises a carbon to nitrogen ratio of 49:1 or greater.

### Claim 16

A method for producing at least one lipid product from a gaseous substrate, the method comprising:
i. receiving a gaseous substrate comprising CO in a first bioreactor containing a culture of at least one microorganism in a liquid nutrient medium, and fermenting the gaseous substrate under anaerobic conditions to produce acetate;
ii. passing at least a portion of the acetate to a second bioreactor containing a culture of at least one microalgae in a liquid nutrient medium, and fermenting the acetate under aerobic conditions to produce at least one lipid product, wherein the microalgae is selected from the group consisting of Thraustochytrium, Japonochytrium, Aplanochytrium, Elina, and Labyrinthula;
iii obtaining a fermentation broth from the second bioreactor;
iv. passing at least a portion of the fermentation broth from the second bioreactor to a separator to produce a product stream comprising at least one lipid and biomass and an acetate depleted permeate stream;
v. passing the permeate stream to an oxygen removal unit to remove oxygen and then recycling at least a portion of the resulting oxygen depleted stream to the first bioreactor.

### Claim 17 (depends on 16)

The method of claim 16, wherein the at least one microorganism in the first bioreactor is selected from the group consisting of Acetobacterium, Moorella, Clostridium, Pyrococcus, Eubacterium, Desulfobacterium, Carboxydothermus Acetogenium, Acetoanaerobium, Butyribacterium, Peptostreptococcus, Ruminococcus, Oxobacter, and Methanosarcina.

### Claim 18 (depends on 16)

The method of claim 16, wherein the at least one microalgae in the second bioreactor is Thraustochytrium.