WO2010098694A2

WO2010098694A2 - Process for production of organic solvents

Info

Publication number: WO2010098694A2
Application number: PCT/RU2010/000053
Authority: WO
Inventors: Evgeniy Rubenovich Davidov; Petr Sergeevich Kanygin; Oleg Anatolievich Frakin; Igor Vladimirovich Cheremnov
Original assignee: "PROF BUSINESS" LLC
Current assignee: "PROF BUSINESS" LLC
Priority date: 2009-02-25
Filing date: 2010-02-09
Publication date: 2010-09-02
Anticipated expiration: 2011-08-25
Also published as: RU2405827C2; RU2009106235A; WO2010098694A3

Abstract

The present invention relates to a process for production of organic solvents, particularly acetone, butanol, ethanol using the anaerobic fermentation by butanol, acetone, and ethanol- producing bacteria. The process comprises saccharifying the pre-milled plant raw material with enzymes which degrade or convert the material into soluble sugars; fermenting the sugars by butanol, acetone, ethanol producing bacteria at a nutrient medium and recovering of organic solvents from the fermentation broth, the process is carried out in one stage, moreover organic solvents are recovered from the fermentation broth by diffuse evaporation through a liquid membrane by applying vacuum or gas.

Description

Process for production of organic solvents

TECHNICAL FIELD OF THE INVENTION

The present invention relates to a process for production of organic solvents, particularly acetone, butanol, ethanol using the anaerobic fermentation by butanol, acetone, and ethanol- producing bacteria.

BACKGROUND OF THE INVENTION

Butanol fermentation, also called acetone butanol ethanol (ABE) fermentation is one of the oldest fermentation processes. Butanol is the most valuable of the produced solvents.

Butanol is an important industrial chemical and is currently used as solvency enhancer in the formation of nitrocellulose lacquers, synthetic resins; as a feedstock chemical in the plastics industry and as a food grade extractant in the food and pharmaceutical industry. As it turned out butanol has excellent fuel characteristics. Compared to the currently popular fuel additive ethanol, butanol is more miscible with gasoline and diesel fuel, has a lower vapor pressure, and is less miscible with water, qualities that make butanol a superior fuel extender than ethanol. Use of butanol as fuel will contribute to clean air by reducing smog-creation compounds, harmful emissions (carbon monoxide).

Currently butanol is produced chemically by either the oxo process starting from propylene (with H₂ and CO over rhodium catalyst or nickel-cobalt catalyst) or the aldol process starting from acetaldehyde.

The recent trend of using butanol as biofuel has revived research efforts aimed at obtaining liquid fuels by anaerobic fermentation.

Now the main directions of ABE process development are focused on metabolic engineering of Clostridium (US 2007259410, WO 2007050671, etc.), improvements in the process productivity and research of new techniques of solvent recovery (US 2005089979, US 5755967).

Traditionally the microbiological process for producing organic solvents from plant material comprises few stages that are performed separately and sequentially. Most processes start with a pretreatment, followed by an enzymatic or chemical hydrolysis (saccharification), a microorganism-based fermentation of the resulting sugars and a recovery of the organic solvents.

For example, WO2008025522 discloses a method of producing bioalcohol, in particular ethanol or butanol, from biomass, the method is carried out sequentially: the biomass is comminuted and saccharified with enzymes, insoluble components and/or non-fermentable sugars are separated off from the biomass, the remaining biomass is fed to a fermentation and the alcohol is obtained from the product of the fermentation.

US 20070178569 discloses a method of making a fuel from biomass material, the method comprising the following separate stages: providing biomass material reduced in size; hydrolyzing the biomass material to provide a hydrolyzed material; combining the hydrolyzed biomass material with Clostridium phytofermentans cells in a medium; and fermenting the hydrolyzed material under conditions and for a time sufficient to produce a fuel.

In the light of the aforesaid there is interest in process for producing organic solvents wherein the sacchariflcation and the fermentation are performed simultaneously, i.e. soluble sugars after saccharification are not accumulated but fermented as they are produced. It allows to optimize the process for producing organic solvents, reduce the operating time, make cheaper by removal of separate reactors and some operations, for example concentration and sterilization of soluble sugars before fermentation.

However there are few obstacles to perform such process. The problem is that the saccharification of the plant raw material and the fermentation of the obtained sugars proceed at the different conditions, temperature, pH. Also it should to be noted that Clostridia are very fastidious (Jones D.T. et al., Acetone-Butanol Fermentation Revisited, MICROBIOLOGICAL REVIEWS, Dec. 1986, Vol. 50, No. 4, p. 484-524).

There is a brief information that researchers at the US Agricultural Research Service (ARS) are refining an integrated method of producing cellulosic biobutanol from wheat straw (Ed./Green Car Congress (2008, October 17) Integrated SSFR Process Could Make Cellulosic Biobutanol More Competitive with Ethanol. [WWW document]. URL http://www.greencarcongress.eom/2008/l Q/integrated-ssfr.html#more). Dilute acid pretreated wheat straw in a bioreactor is saccharified by a combination of enzymes and fermented by a culture of C. beijerinckii P260, which produces a combination of acetone, biobutanol and ethanol (ABE). However the reference does not disclose the details of this process, either the possibility of continuous mode. The researchers indicated the drawback of the process. In early trial runs the pace of fermentation outran the pace of hydrolysis and small batches of additional sugar were added in the process. Also the pretreatment with dilute acid refers to the drawback because it causes known difficulties of process structure.

It should to be noted that the main drawbacks of the fermentation process relate to techniques of solvent recovery from the fermentation broth. The separation of the organic solvents from the fermentation broths is generally performed by distillation, but this procedure is energy-intensive and may adversely affect the economics of the fermentation process, particularly when dilute solutions are involved. The production of alcohols by fermentation invariably presents a dilute solution for product recovery, since the producing organism is subject to product inhibition at low concentrations of the organic solvents.

Many attempts have been made to economize solvent recovery. The product removal techniques, including adsorption, liquid— liquid extraction, perstraction and gas stripping, have been developed to reduce the cost of butanol recovery (Food biotechnology, Second Edition, edited by Kalidas Shetty, et al., CRC Press, 2006, p.527).

Recovery processes which involve the use of membranes for separating organic solvents from fermentation broth have been used increasingly in recent years.

Particularly US 5755967 discloses a pervaporation process for the selective removal of acetone and/or butanol from an aqueous solution comprising acetone or butanol, or mixtures thereof. The silicalite, silicalite filled polymer membrane is suited to the removal of acetone and butanol from Clostridium acetobutylicum fermentation media.

The use of a liquid membrane, rather than a solid membrane, has been described by Matsumura (abstract, Matsumura et al., Energy saving effect of pervaporation using oleyl alcohol liquid medium in butanol purification, Bioprocess Engineering 3, 93-100, 1988). The oleyl alcohol containing liquid membrane was supported on a microporous 25 mm thick flat polypropylene sheet. It was estimated that if this pervaporation membrane was used for butanol separation, the energy requirements would be only 10% of that of conventional distillation. Unfortunately this was not a stable membrane.

Summarizing the aforesaid, a need exists for an improved continuous process to produce organic solvents from non-edible renewable plant material, particularly forestry residues, wherein the saccharification, the fermentation and the recovery of the organic solvents from the fermentation broth are performed simultaneously, preferably in one stage and in a single vessel.

BRIEF SUMMARY OF THE INVENTION

In accordance with the present invention, there is provided a process for production of organic solvents comprising:

-saccharifying the pre-milled plant raw material with enzymes which degrade or convert the material into soluble sugars;

-fermenting the sugars by butanol, acetone, ethanol producing bacteria at a nutrient medium and -recovering of organic solvents from the fermentation broth, wherein the process is carried out in one stage, moreover organic solvents are recovered from the fermentation broth by diffuse evaporation through a liquid membrane by applying vacuum or gas.

Preferably the saccharification and the fermentation are carried out in a single vessel.

In one embodiment the liquid membrane is a layer of hydrophobic liquid on the surface of the fermentation broth. The hydrophobic liquid is synthetic and/or vegetable oil. For example, the synthetic oil is silicone oil. The thickness of the layer of the hydrophobic liquid is about 3-30 mm.

In one embodiment an even rate is maintained for the saccharification and fermentation. The sugar concentration and the concentration of solvents are monitored to maintain the even rate. The sugar concentration in the fermentor is maintained at between about 1.2-2.5%. The process is carried out at pH 4.2-5.0 and temperature 33-55 ⁰C.

In one embodiment, plant material is wood chips, hi one embodiment, the wood chips are coniferous wood chips. In the case the process further comprises removal of pitch, wherein removal of pitch is carried out by extraction with organic solvents. Preferably enzyme complex is used for saccharification. The enzyme complex is matched to polysaccharide components of the raw plant material, hi one embodiment, the culture liquid obtained by cultivation (growth) of fungi Penicillium verruculosum is used as the enzyme complex, hi one embodiment, butanol, acetone, ethanol producing bacteria are Clostridium acetobutylicum.

DETAILED DESCRIPTION OF THE INVENTION hi accordance with the present invention plant material includes various agricultural residues (straws, hulls, stems, stalks); deciduous and coniferous woods. One of advantages of the present invention is processing of forestry residues, particularly wood chips.

In accordance with the present invention the pre-milling of the plant material includes the coarse milling and the fine milling. The coarse milling of the raw material is carried out in mills, preferably in ball-mills to particles ranging in size from 1 mm to 2 mm. The operation gives option to dry the raw material and prepare it for the fine milling. The fine milling is carried out to particles ranging in size from 1 μm to 5 μm. hi one preferred embodiment, the plant material is coniferous wood chips, hi this case the removal of pitch is preferable before the fine milling; it gives a possibility to carry out the complete saccharification of cellulose and hemicellulose otherwise the part of the enzymes are sorbed by pitch and do not participate hi the process. Wood chips are subjected to a solvent extraction process, preferably with acetone or ethanol (plant material: solvent ratio is 1:7 - 1:10). Preferably enzyme complex is used for saccharifϊcation. The enzyme complex is a complex of cellulases, cellobiases, xylanases and/or mannanases. The enzyme complex is matched to polysaccharide components of the raw plant material, e.g. it depends on the sort of wood chips (hard or soft). It is possible to use the culture liquid obtained by cultivation (growth) of strains of fungi Penicillium verruculosum, for example VKM F-3984D, as the enzyme complex.

Butanol, acetone, ethanol producing bacteria includes species of Clostridium, including Clostridium beijerinckii and Clostridium acetobutylicum, as well as another bacteria known in the art.

In accordance with the present invention the saccharifϊcation, the fermentation and recovery of the organic solvents from the fermentation broth are performed in one stage, hi the preferred embodiment the processes are carried out in a single vessel. The number of vessels is reduced resulting in capital cost savings. Furthermore, the presence of alcohol during saccharification reduces the likelihood of contamination, especially in continuous operations.

The following conditions are optimal for such one-stage process: pH 4.2-5.0 and temperature 33-55 ⁰C. Also an even rate is maintained for the saccharification and fermentation. The sugar concentration and the concentration of solvents are monitored to maintain the even rate for the saccharification and the fermentation. The sugar concentration in the fermentor is maintained at between about 1.2-2.5%. When the sugar concentration in the fermentor is reduced, the freshly-mixed suspension of the wood powder in water and the enzyme complex are added.

It should to be noted that the claimed process in comparison to the processes from the prior of art allows that the amount of sugar that comes into solution per time unit for the sachharification is approximate to the amount of sugar consumed by Clostridium.

Recovery of organic solvents is achieved by diffuse evaporation through a liquid membrane by applying vacuum or gas. In one embodiment the liquid membrane is a layer of hydrophobic liquid on the surface of the fermentation broth. Therefore the hydrophobic liquid, for example silicone oil, is placed in a fermentor before the fermentation process. When the solvent concentration in the fermentor is approaching to inhibitory value for the producing culture, the overhead pressure in the fermentor is reduced and the produced solvents are removed through the layer of the hydrophobic liquid, then the solvents are condensed in a cooling unit.

The invention may be further understood by the following non-limiting examples.

Example 1 The wood powder with particle size of l-5μm is suspended in water and the complex of hydrolytic enzyme (cellulase, xylanase and/or mannanase, cellobiase) is added to the suspension in the ratio 2.5 g per 1 kg of the fined wood powder. The saccharification was allowed to proceed at 55 ⁰C, pH 5.5 and in 12 hours the process is completed. 45% of carbohydrates are in the solution and lignin is removed from the sugar solution by centrifugation.

Obtained 2.7% carbohydrate solution was sterilized and then transferred into a fermentor. Then the mixture was inoculated with inoculum of Clostridium acetobutylicum VKM B-2531D with density of 1-2 milliard/ml. The fermentation was allowed to proceed at 37 ⁰C, pH 3.8 for 48 hours.

Then the vacuum of -0.4 atm abs was applied, the solvent vapors were taken off. The ABE vapors were cooled in a condenser and in result solution, containing butanol, acetone and ethanol was obtained (ABE ratio is 10:88:2).

After removal of the solvents and the fermentation gas under vacuum, the nutrient medium, containing 75 g of sugars is added per day with the pace of 100 ml per hour. One time in two days 28 g of yeast autolyzate is added into the fermentor. Productivity was 8 g/l/day. Solvent yield on sugars is 27-33%. Sugar yield on the used wood powder is 14-16%.

Example 2

Wood chips, e.g. coniferous wood chips are milled to size of 1-2 mm. Then the obtained wood particulates are extracted with acetone for pitch removal. The extracted wood particulates are milled to the size of l-5μm. The obtained wood fine powder is mixed with water (density lg/cm³). The suspension is charged into a fermentor. hi the fermentor the complex of hydrolytic enzyme (cellulase, xylanase and/or mannanase, cellobiase) is added (ratio 2.5 g per 1 kg of the milled wood chips). Strains of Penicillium verruculosum (for example, VKM F-3984D) are a source of cellulases and hemicellulases, as well as other enzymes useful in the enzymatic hydrolysis of major polysaccharides so the culture liquid obtained by cultivation of the fungi can be used for saccharifying instead the complex.

The standard mixture of mineral salts, vitamins was added into the fermentor. Then the medium was inoculated with cells of Clostridium acetobutylicum (e.g. VKM B-2531D) with density of 1-2 milliard/ml. Silicone oil was layered on the liquid surface; thickness of layer was 10 mm. The saccharification and fermentation were allowed to proceed at 37 ⁰C, pH 4.2-4.5. The sugar concentration was 2%. If the sugar concentration in the fermentor is reduced, the freshly-mixed suspension of the wood powder in water and the enzyme complex are added. For butanol molecules having the diphilic properties, the minimum energy state is achieved by orientation of these molecules as follows: polar hydroxy 1 groups orient to the water phase with their fatty hydrocarbon chains oriented towards the layer of oleic acid.

When the concentration of solvents in the feπnentor approached the following values: 1.5 % of butanol, 0.7% of acetone and 0.2% of ethanol, the vacuum of -0.4 atm abs was applied. The reduced pressure provides evaporation of the ABE vapors. The removed gases are directed into cooling device wherein the solvents are condensed and collected in the accumulator vessel.

The layer of silicone oil keeps out the transfer of water into cooling device.

The vapors are removed and directed to the vapor condenser. 75 ml of the aqueous solution (64 ml of organic solvents and 11 ml of water; the presence of water results from that the fermentation gases pass through membrane and water is directed into the cooling device with gases), contained 60% butanol, 20% acetone and 5% ethanol, was obtained.

In should to be noted that the vacuum removal of the organic solvents without the liquid membrane results in that the water vapors are fallen into the cooling unit and the butanol concentration is no more that 10% in the aqueous solution.

After removal of the solvents and the fermentation gases under vacuum, the nutrient medium, containing freshly-mixed suspension of the wood powder in water, enzyme complex are added with the pace of 100 ml per hour. One time in two days 28 g of yeast autolyzate is added into the fermentor. Productivity was 8 g/l/day. Solvent yield on sugars is 27-33%. Sugar yield on the used wood powder is 14-16%.

Therefore the present invention provides the improved continuous process to produce organic solvents from non-edible renewable plant material, particularly forestry residues, wherein the saccharification, the fermentation and the recovery of the organic solvents from the fermentation broth are performed simultaneously, preferably in one stage and in a single vessel.

Claims

1. A process for production of organic solvents comprising:

-fermenting the sugars by butanol, acetone, ethanol producing bacteria at a nutrient medium and

-recovering of organic solvents from the fermentation broth, wherein the process is carried out in one stage, moreover organic solvents are recovered from the fermentation broth by diffuse evaporation through a liquid membrane by applying vacuum or gas.

2. The process of claim 1 wherein the liquid membrane is a layer of hydrophobic liquid on the surface of the fermentation broth.

3. The process of claim 2 wherein the hydrophobic liquid is synthetic and/or vegetable oil.

4. The process of claim 3 wherein the synthetic oil is silicone oil.

5. The process of claim 2 wherein the thickness of the layer of the hydrophobic liquid is about 3-30 mm.

6. The process of claim 1 wherein an even rate is maintained for the saccharification and fermentation.

7. The process of claim 6 wherein the sugar concentration and the concentration of solvents are monitored to maintain the even rate.

8. The process of claim 1 wherein the sugar concentration in the fermentation broth is maintained at between about 1.2-2.5%.

9. The process of claim 1 wherein the saccharification and the fermentation are carried out at pH 4.2-5.0.

10. The process of claim 1 wherein the saccharification and the fermentation are carried out at the temperature of 33-55 ⁰C.

11. The process of claim 1 wherein the plant raw material is wood chips.

12. The process of claim 11 wherein the wood chips are coniferous wood chips.

13. The process of claim 12, further comprising extraction of pitch.

14. The process of claim 1 wherein an enzyme complex is used for the saccharification and the complex is matched to polysaccharide components of the plant material.

15. The process of claim 14 wherein the culture liquid obtained by cultivation of Penicillium verruculosum is used as the enzyme complex.

16. The process of claim 1 wherein said butanol, acetone, ethanol producing bacteria are Clostridium acetobutylicum.