Process for the manufacture of butanol or acetone

Abstract

A process for the manufacture of butanol, acetone and/or other renewable chemicals is provided wherein the process utilises one or more of the group comprising by-products of the manufacture of malt whisky, such as draff, pot ale and/or spent lees, biomass substrates, such as paper, sludge from paper manufacture and spent grains from distillers and brewers, and diluents, such as water and spent liquid from other fermentations. The process comprises treating a substrate to hydrolyse it and fermenting the treated substrate at an initial pH in the range of 5.0 to 6.0. Also provided is a biofuel comprising butanol manufactured according to the process of the invention.

Claims

1. A process for the manufacture of butanol and/or acetone, comprising at least the steps of: obtaining pot ale, the pot ale being a byproduct of spirit distillation for production of malt whiskey, wherein the pot ale is a distillation residue from a copper distillation vessel comprising a distillable wash from a yeast fermentation of grain comprising malted barley; treating a carbohydrate substrate comprising draff, with the pot ale, to provide a treated substrate, said draff comprising spent grain consisting essentially of malted barley; and fermenting the treated substrate in the presence of a culture of butanol- and/or acetone-forming micro-organisms at an initial pH in the range of 5.0 to 6.0 and at a concentration of free copper ions of less than 20 μM to provide a fermented product containing butanol and/or acetone.

2. The process of claim 1, wherein the carbohydrate substrate comprises non-starch components of malted barley, said non-starch components being digestible fiber, protein and oil from the malted barley.

3. The process of claim 1, wherein the carbohydrate substrate consists essentially of draff and is treated with an acid to hydrolyze the carbohydrate substrate consisting essentially of draff.

4. The process of claim 1, wherein the carbohydrate substrate consists essentially of draff and is treated with one or more enzymes to hydrolyze the carbohydrate substrate consisting essentially of draff.

5. The process of claim 1, wherein the carbohydrate substrate consists essentially of draff and is treated with both an acid and one or more enzymes to hydrolyze the carbohydrate substrate consisting essentially of draff.

6. The process as claimed in claim 1, wherein the step of treating the substrate further comprises: hydrolysing the carbohydrate source in the presence of water and hydrogen ions or water and hydroxide ions.

7. The process as claimed in claim 1, wherein the step of treating the substrate further comprises: hydrolysing the carbohydrate source in the presence of an aqueous solution of sulphuric acid.

8. The process as claimed in claim 1, wherein the step of treating of the substrate further comprises: treating the carbohydrate source with one or more enzymes.

9. A process for the manufacture of butanol and/or acetone, comprising at least the steps of: obtaining pot ale from a malt whisky distillery, pot ale being a by-product of a first spirit distillation, wherein the pot ale is a distillation liquor residue from a copper distillation vessel containing a distillable wash from a yeast fermentation of grain comprising malted barley; treating a carbohydrate substrate comprising draff with the pot ale, to provide a pot ale-treated carbohydrate substrate; and fermenting the pot ale-treated substrate in the presence of a culture of butanol- and/or acetone-forming micro-organisms at an initial pH in the range of 5.0 to 6.0 and at a concentration of free copper ions of less than 20 μM to provide a fermentation product containing butanol and/or acetone.

10. The process of claim 9, wherein the carbohydrate substrate comprises non-starch components of malted barley, said non-starch components being digestible fiber, protein and oil from the malted barley.

11. The process of claim 9, wherein the carbohydrate substrate consists essentially of draff and is treated with an acid to hydrolyze the carbohydrate substrate consisting essentially of draff.

12. The process of claim 9, wherein the carbohydrate substrate consists essentially of draff and is treated with one or more enzymes to hydrolyze the carbohydrate substrate consisting essentially of draff.

13. The process of claim 9, wherein the carbohydrate substrate consists essentially of draff and is treated with both an acid and one or more enzymes to hydrolyze the carbohydrate substrate consisting essentially of draff.

14. The process as claimed in claim 9, wherein the step of treating the substrate further comprises: hydrolysing the carbohydrate source in the presence of water and hydrogen ions or water and hydroxide ions.

15. The process as claimed in claim 9, wherein the step of treating the substrate further comprises: hydrolysing the carbohydrate source in the presence of an aqueous solution of sulphuric acid.

16. The process as claimed in claim 9, wherein the step of treating of the substrate further comprises: treating the carbohydrate source with one or more enzymes.

Description

BRIEF DESCRIPTION OF THE FIGURES

(1) FIG. 1 shows the influence of initial pH on fermentation of acid and enzyme pre-treated draff in pot ale by C. acetobutylicum ATCC 824. Draff was pre-treated with 0.08 M H.sub.2SO.sub.4 and the pH adjusted to between pH 5.0-6.0 prior to enzyme addition. After enzyme hydrolysis, the pH was adjusted to 4.5, 4.8, 5.0, 5.5, 6.0 or 6.5 for fermentation. FIG. 1(a) shows sugars resulting from acid and enzyme treatment, FIG. 1(b) shows residual sugars after fermentation, FIG. 1(c) shows the ABE products from fermentation and FIG. 1(d) shows yield of butanol and ABE from draff;

(2) FIG. 2 compares ABE production by C. acetobutylicum ATCC 824 from acid-pre-treated draff in either water or pot ale. After acid treatment, the pH was adjusted to pH 5.5 and enzymes and microorganisms added;

(3) FIG. 3 shows ABE production by C. acetobutylicum ATCC 824 and C. beijerinckii NCIMB 8052 from draff at 1 L scale; and

(4) FIG. 4 shows ABE production by C. saccharoperbutylacetonicum NCIMB 12606 from (a) white office paper and (b) newspaper dissolved in either water or 50% pot ale.

EXAMPLES

(5) General Methods

(6) The following organisms were used: C. acetobutylicum ATCC 824, C. beijerinckii NCIMB 8052 and C. saccharoperbutylacetonicum NCNB 12606. Clostridia were maintained as spore suspensions at 4° C. Spores were heat shocked at 80° C. for 10 minutes and inoculated into reinforced clostridia media (RCM, Oxoid Ltd, Cambridge, UK). Cultures were incubated for 24 hours and then subcultured into tryptone-yeast extract-ammonium acetate media (TYA) media containing glucose before being used as a starting culture (at 5% v/v) for all experiments. TYA consisted of (g/l) tryptone, 6; yeast extract, 2; ammonium acetate, 3; KH.sub.2PO.sub.4, 0.5; MgSO.sub.4.Math.7H.sub.2O, 0.3; FeSO.sub.4.Math.7H.sub.2O, 0.01 supplemented with 5% glucose. All clostridia cultures were incubated in an anaerobic workstation under an N.sub.2—H.sub.2—CO.sub.2 (80:10:10) atmosphere at 33° C. For 1 L scale, fermentations were conducted in fermenters (Biostat A PILES, Sartorius Stedim Ltd, Surrey, UK). Oxygen-free conditions were achieved by sparging the media in the fermenters with oxygen-free N.sub.2 for 1 hour prior to inoculation with clostridia. For all 1 L fermentations, agitation was set at 200 rpm and temperature at 33° C.

(7) Wet draff as received from the distilleries, had a moisture content between 75-80%. Where stated, draff was dried at 80° C. to a moisture content of approximately 4% and milled prior to further processing.

(8) Solvents (ethanol, acetone and butanol) were analysed using a Chrompack 9001 gas chromatograph equipped with a flame ionisation detector and a CP SIL 5CB column of length 10 m and diameter 0.32 mm (all Chrompack, Middelburg, Netherlands). All samples were filtered through 0.2 μm cellulose acetate syringe filters before analysis and concentrations were determined by reference ethanol, acetone and butanol standards.

(9) For acid (acetic and butyric) and monosaccharide (glucose, xylose and arabinose) analysis, samples were filtered through 0.2 μm syringe filters and acidified with H.sub.2SO.sub.4. Samples were analysed by HPLC using a Varian 920 LC fitted with integrated UV-VIS dual wavelength and refractive index detectors (Varian Ltd., Oxford, UK). Components were separated at room temperature on a Rezex ROA Organic acid H.sup.+ 8% 300×7.8 mm column (Phenomenex, Cheshire, UK) with 0.005 N H.sub.2SO.sub.4 as the mobile phase at a flowrate of 0.5 ml/min. Acids were detected at 210 nm while sugars were detected with the RI detector and concentrations were determined by reference to the corresponding standards.

EXAMPLE 1

Composition of Draff

(10) Draff was collected from three different malt distilleries in Scotland. The monosaccharide composition of the draff was analysed according to the Laboratory Analytical Procedure developed by the National Renewable Energy Lab for the analysis of structural carbohydrates (Sluiter et al., 2008. NREL. Laboratory analytical procedure for the determination of structural carbohydrates and lignin in biomass. NREL/TP-510-42618). The results of the analysis are provided in Table 1. Glucose, xylose and arabinose were the predominant sugars, with very low levels of galactose (less than 2%) and no mannose detected. There was little variation in the sugar composition of draff from different distilleries. Based on these values, complete hydrolysis of draff (10.5% dry draff (w/v) as used in the experiments detailed below) should yield approximately 50 g/l monosaccharide.

(11) TABLE-US-00001 TABLE 1 Monosaccharide composition of draft Sugar (g/100 g draff) Source Glucose Xylose Arabinose Total Distillery 1 20.9 ± 0.2 21.3 ± 0.1 9.0 ± 0.2 51.2 ± 0.2 Distillery 2 18.4 ± 0.2 21.3 ± 0.2 9.2 ± 0.0 48.8 ± 0.4 Distillery 3 20.5 ± 0.0 21.6 ± 0.3 9.3 ± 0.0 51.3 ± 0.3

EXAMPLE 2

Effect of pH control on solvent production by clostridia

(12) The effect of pH on fermentation of glucose in TYA media by C. acetobutylicum ATCC 824 was investigated. Fermentations were conducted at 1 L scale and the pH was controlled at a range of set points between pH 4.5-6.5 with automated addition of either alkali or acid, At pH 4.5, no glucose utilisation, acid or ABE production was detected. For all other fermentations, glucose was completely consumed within 48 hours and acids (butyric and acetic) and solvent (acetone, butanol and ethanol) were produced (Table 2). ABE production was highest at pH 4.8 and 5.0, corresponding to yields of 0.34 and 0.30 g ABE/g sugar, respectively. Acid production increased between pH 5.5 to 6.5, with a corresponding decrease in conversion of sugar to ABE. At pH 6.5, acids only were produced with final concentrations of 7.8 and 12.8 g/l acetic and butyric acid, respectively.

(13) TABLE-US-00002 TABLE 2 Conversion of 5% glucose to acid and ABE by C. acetobutylicum ATCC 824 in TYA media controlled at either pH 4.8, 5.0, 5.5, 6.0 or 6.5. Acid (butyric and acetic) and ABE concentrations were determined after 68 hours with ABE yield expressed as g of ABE produced per g of sugar consumed. pH Acid (g/l) ABE (g/l) Yield (g ABE/g sugar) 4.8 0.7 15.2 0.34 5.0 0.9 14.3 0.30 5.5 7.9 12.3 0.25 6.0 13.6 6.7 0.13 6.5 20.5 0.6 0.01

EXAMPLE 3

Pot Ale as a Growth Medium

(14) Pot ale was collected from a Scottish malt distillery and analysed for copper content. The pot ale had 71.8 μM total Cu of which 21.1 μM was determined to be available as “free” Cu in the supernatant with the rest bound to the solids. To assess whether this Cu concentration was toxic to C. acetobutylicum ATCC 824, fermentation of 5% glucose in 100 ml TYA media supplemented with different concentrations of Cu was compared (Table 3). Cu had no effect on ABE production at 5 and 10 μM with ABE concentrations of approximately 12 g/l being similar to that of the control without Cu. At the higher Cu concentration, ABE concentration was reduced to 8.6 g/l, indicating that at this concentration Cu was inhibitory to clostridia. As the pot ale had a “free” Cu content of 21.1 μM, it was decided to test clostridia fermentation in half strength pot ale in order to reduce the Cu concentration below inhibitory levels. Half-strength pot ale supplemented with glucose provided enough nutrients for growth of 824 with ABE production similar to the TYA control (Table 3),

(15) TABLE-US-00003 TABLE 3 Conversion of 5% glucose to ABE by C. acetobutylicum ATCC 824 in either TYA, TYA containing 5, 10 or 20 μM Cu or 50% pot ale. Media ABE (g/l) TYA 12.4 ± 0.3 TYA, 5 μM Cu 12.3 ± 0.3 TYA, 10 μM Cu 11.6 ± 0.1 TYA 20 μM Cu  8.6 ± 2.0 50% pot ale 12.0 ± 1.7

EXAMPLE 4

Influence of Initial pH on Fermentation of Hydrolysed Draff

(16) The effect of initial pH on fermentation of pre-treated draff was investigated. Dried, milled draff was pre-treated by adding 10.5% (w/v) to 250 ml duran bottles with 0.08 M H.sub.2SO.sub.4 in 50% pot ale and sterilised at 121° C. for 15 min. After cooling, the pH was adjusted to between pH 5.0-6.0 by addition of 10 M NaOH and incubated with cellulase and hemicellulase enzymes at 33° C. for 24 hours. For fermentation, the pH of the solutions was adjusted to either 4.5, 4.8, 5.0, 5.5, 6.0 or 6.5 prior to inoculation with C. acetobutylicum ATCC 824. The initial sugar concentration was monitored before fermentation and the residual sugar, ABE concentration and ABE yield were calculated after fermentation (FIG. 1). The initial concentration of sugars was similar for all samples, with approximately 9.6, 11.2, and 9.9 g/l glucose, xylose and arabinose. No growth or gas production was apparent at pH 5.0 or lower and no sugars were utilised. ABE production was greatest at pH 5.5 (14.2 g/l) with a yield of 13.2 g/100 g draff. This was reduced at pH 6.0, with 9.3 g ABE/100 g draff At pH 6.5, approximately half the sugar was utilised but there was poor conversion to ABE with a final concentration of 2.3 g/l.

EXAMPLE 5

Fermentation of Add Pre-treated Draff in Pot Ale or Water

(17) Dried, milled draff (10.5% w/v) was pre-treated with 0.08 M H.sub.2SO.sub.4 in either water or pot ale in 250 ml duran bottles by sterilisation at 121° C. for 15 minutes. After cooling the pH was adjusted to 5.5 by the addition of 10 M NaOH, Cellulase and hemicellulase enzymes and C. acetobutylicum ATCC 824 inoculum were added and bottles incubated at 33° C. The ABE concentration was determined after fermentation (FIG. 2). For draff in water, ABE yield was 14.0 g ABE/100 g draff whereas in pot ale, a yield of 14.9 g ABE/100 g draff resulted.

EXAMPLE 6

Conversion of Draff to Butanol and Acetone at 1 L Scale

(18) Draff (10.5% w/v) was pre-treated with 0.08 M H.sub.2SO.sub.4 in 50% pot ale in 1 L fermenters by sterilisation at 121° C. for 15 minutes. In this case draff was used wet, as received from the distillery, without any further processing. After cooling to 33° C., the pH was adjusted to pH 5.5 by the addition of 10 M NaOH and the fermenters were sparged with N.sub.2. After degassing, enzymes and either 824 or 8052 were added and solvents were analysed at the end of the fermentation. Fermentation by C. acetobutylicum ATCC 824 and C. beijerinckii NCIMB 8052 resulted in ABE levels of 11.3 and 12.8 g/l, respectively (FIG. 3). This corresponded to conversion rates of 10.6 and 12.1 g ABE per 100 g draff, respectively.

EXAMPLE 7

Process for Conversion of Waste Paper to Butanol and Acetone

(19) White office paper and newspaper were shredded to 5 mm wide strips and 6.7% (w/v) was mixed with either water or 50% pot ale in 250 ml duran bottles and the pH adjusted to pH 5.5. After sterilisation, the bottles were cooled and cellulase and C. saccharoperbutylacetonicum NCIMB 12606 added. After fermentation, the ABE concentrations were determined (FIG. 4). There was poor conversion of paper to ABE in water compared to pot ale, demonstrating that pot ale was required to provide additional nutrients. In pot ale, the ABE yields after fermentation with C. saccharoperbutyfacetonicum were 24.8 g ABE per 100 g office paper and 16.8 g ABE per 100 g newspaper.

(20) Although the invention has been described in connection with specific preferred embodiments, it should be understood that the invention as claimed should not be unduly limited to such specific embodiments. Indeed, various modifications of the described modes of carrying out the invention which are obvious to those skiiled in the art are intended to be covered by the present invention.