CN104357491B - The preprocess method of butanol is produced in a kind of utilization bagasse fermentation - Google Patents
The preprocess method of butanol is produced in a kind of utilization bagasse fermentation Download PDFInfo
- Publication number
- CN104357491B CN104357491B CN201410744063.8A CN201410744063A CN104357491B CN 104357491 B CN104357491 B CN 104357491B CN 201410744063 A CN201410744063 A CN 201410744063A CN 104357491 B CN104357491 B CN 104357491B
- Authority
- CN
- China
- Prior art keywords
- filtrate
- solid residue
- afterwards
- fermentation
- iii
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P7/00—Preparation of oxygen-containing organic compounds
- C12P7/02—Preparation of oxygen-containing organic compounds containing a hydroxy group
- C12P7/04—Preparation of oxygen-containing organic compounds containing a hydroxy group acyclic
- C12P7/16—Butanols
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02E—REDUCTION OF GREENHOUSE GAS [GHG] EMISSIONS, RELATED TO ENERGY GENERATION, TRANSMISSION OR DISTRIBUTION
- Y02E50/00—Technologies for the production of fuel of non-fossil origin
- Y02E50/10—Biofuels, e.g. bio-diesel
Abstract
The invention discloses the preprocess method that butanol is produced in a kind of fermentation of utilization bagasse, the method includes mechanical crushing, liquid hot water treatment, microwave treatment, microorganism decomposition, ammonia treatment and enzymolysis processing step, substep diastatic fermentation and simultaneous saccharification and fermentation are carried out by the catabolite that each step after above-mentioned steps, can be merged.The preprocess method by the control of the content of major inhibitors furtural and hydroxymethylfurfural below the concentration of the bacteria growing that do not influence to ferment, and can not produce other inhibitor such as glucuronic acid, p-Coumaric Acid, syringic acid and forulic acid substantially.The method just can directly carry out biofermentation and produce butanol without carrying out detoxification purification process to pretreatment catabolite, have the advantages that controllable inhibitor generation, low cost, yield are high and environment-friendly.
Description
Technical field
The present invention relates to living beings preprocessing technical field, and in particular to is produced from the pretreatment side of butanol in a kind of utilization bagasse fermentation
Method.
Background technology
At present, it is existing various ripe to be directed to fibrous material and the preprocess method of half fibrous material obtains sugar to hydrolyze
Point, the sugar of gained can be used for fermenting and producing bio-fuel.
Recently, these methods obtained some progress in industrial biotechnology field, were cost-effective to be produced using agricultural
Industry waste material provides potential chance.For example, bagasse is a kind of comprehensive material, it is the Main By product in Sugarcane Industry,
It is also one of Main Agricultural waste material, all richness originates in southern china every year.Bagasse contains about 50% cellulose, the half of 25% fibre
Dimension element and 25% lignin, with its produce biological butanol be a kind of Perfected process for producing bio-fuel because butanol compared to
Traditional ethanol, with calorific value and lower hygroscopicity higher.
However, it is whether feasible using bagasse production butanol, depend on including many factors such as pretreatment and detoxification purifying.And
Traditional preprocess method, such as acidolysis, often produce the serious toxicant for suppressing clostridium growth.
Many researchs report the preprocess method of various cereal residues that can be used for fibre-rich element and hemicellulose, bag
Include steam explosion or dilute acid pretreatment method.Acid hydrolyzation is a kind of conventional and effective preprocess method, and particularly common is dilute
Sulfuric acid.The method is not only simple, and with low cost.However, in acidolysis, often producing a quasi-microorganism inhibitor, the suppression
Agent can suppress the growth of microorganism in fermentation.Furtural and hydroxymethylfurfural are the main suppression produced in biofermentation
Agent.Therefore, before biological butanol fermentation is carried out, hydrolysate must first carry out detoxification purifying, to remove inhibitor.
However, pretreated detoxification purifying is a process for complexity, although various detoxification purification process have been obtained
Research, such as lime treatment, evaporation, ion-exchange chromatography method, activated carbon and biological treatment, otherwise but these methods
Detoxification purification effect is bad, otherwise cost is too high.At present, main current method is to remove inhibitor using chemical method, because
Chemical method is relatively simple, and cost is relatively low.However, during using chemical method, due to the otherness of the chemical constitution of inhibitor, causing
The method cannot be purged to all inhibitor.In addition, chemical method can also produce many salts, cause to contain height in zymotic fluid
The salt of concentration, and high salt concentration can produce serious inhibition to microbial fermentation, so as to cause subtracting for biological butanol yield
It is few.
Therefore, suitable preprocess method is selected just to seem particularly significant.During selection preprocess method, following two need to be considered
Point：1st, the maximally effective destruction fibre structure of energy, to produce the sugar of abundance, and prevents the loss to sugar, while being also avoided that or subtracting
The generation of few inhibitor；2nd, preprocess method need to be with low cost, simple and easy to apply.
In the prior art, not having can also avoid or reduce the preprocess method that all major inhibitors are produced.Therefore, urgently
A kind of both economically feasibles to be developed, and all major inhibitors contents can be controlled, produce butanol to carry out fermentation to bagasse
Preprocess method.
The content of the invention
It is an object of the invention to provide the preprocess method that butanol is produced in a kind of fermentation of utilization bagasse, the preprocess method
The generation of toxin for suppressing growth of microorganism can be controlled.By the generation for controlling toxin so that bagasse after pretreatment
Without follow-up purifying detoxification treatment, fermenting and producing butanol just can be carried out, greatly reduce production cost.
The preprocess method comprises the following steps：
1）Mechanical crushing：It is 5-10cm that segment to length is carried out to sugarcane, is dried 2 days at 65 ± 2 DEG C, then carries out powder
It is broken, 20 mesh sieves are crossed, obtain bagasse powder of the particle diameter less than 830 μm；
2）Liquid hot water treatment：It is 250g/L by solid-liquid mass volume ratio, by step 1）Gains under normal temperature in water
Immersion 12h, preheats 45s with steam afterwards, is subsequently adding liquid hot water, at 200 ± 3 DEG C, reacts 1h, is carried out after reaction
Filter, collects filtrate I and solid residue I；
3）Microwave treatment：Set-up procedure 2）The humidity of gained solid residue I is 40%, is placed in reactor, Ran Houtong
Enter nitrogen and exclude oxygen, in 8-12h is placed at 4 DEG C, reactor is placed in microwave-heating stove afterwards, 7 are reacted at 120 DEG C
Min, then reaction gains are cooled to normal temperature, reaction gains are embathed with 90% ethanol, filtered afterwards, collect
Filtrate II and solid residue II；
4）Microorganism is decomposed：Baking step 3）Gained solid residue II, adds nutrient solution to be mixed afterwards, then at
Sterilized at 121 DEG C, after sterilizing, it is 30% to adjust its humidity, by percentage to the quality, adds 10% decomposer bacterium solution,
In 10 d that fermented at 30 DEG C, filtered afterwards, collected filtrate III and solid residue III；
The formula of the nutrient solution is：30 mg H3BO3、20 mg MnCl2.4H2O、185 mg ZnSO4•7H2O、20
mg Na2MoO4•2H2O、280 mg FeSO4•H2O、200 mg CuSO4、5 g NH4NO3、1 g KH2PO4、0.15 g MgSO4•
H2O、0.11 g CaCl2With 1 L ddH2O, pH are 7.0；
The decomposer isAspergillus niger ATCC 1015、Trichoderma reesei ATCC 26921
WithPenicillium janthinellum One kind in ATCC 44750；
5）Ammonia treatment：Use ddH2O is to step 4）Gained solid residue III is cleaned, and is baked to afterwards, with solid-liquid
Mass ratio is 1:2, the solid residue III after drying is mixed with ammoniacal liquor, under 0.1MPa pressure, 30 are reacted in 90 DEG C
Min, is filtered afterwards, collects filtrate IV and solid residue IV；
6）Enzymolysis processing：The solid residue IV is soaked into 2 h in water, regulation pH is 7.0, is dried after concentration
It is dry, then prepare enzymolysis mixture, every 500 ml enzymolysis mixtures are made up of following substances：The solid that 50 g have been dried
Residue IV, 10g cellulases, 100 ml OPTMASH BG, 5 mg chloramphenicol and 2.5 mg kanamycins；Adjustment enzymolysis is mixed
The pH of compound is 5.0, is 55 DEG C in temperature, and shaking speed is filtered afterwards under 220rpm, to react 60 h, collects filtrate
V and solid residue V；
7）Filtrate I, filtrate II, filtrate III, filtrate IV and solid residue IV are merged, SSF fermentations are remained, or will be described
Filtrate I, filtrate II, filtrate III, filtrate IV and filtrate V merge and concentrated by rotary evaporation, remain SHF fermentations.
Preferably, when microorganism decomposition is carried out, decomposer used isTrichoderma reesei ATCC 26921。
In existing pretreatment mode, when a certain pretreatment mode is used alone, it is impossible to bagasse fibre of fully degrading
Element, sugar yield is extremely low；Such as it is combined with diluted a cid method, although sugar yield makes moderate progress, but corrosion can be produced to reactor and is occurred dirty
Dye problem, meanwhile, the control effect to inhibitor concentration is also undesirable.
The present invention selects hot water treatment high first, first passes through and soaks 12h in water at normal temperatures, is preheated with steam afterwards
45s, is subsequently adding liquid hot water, at 200 ± 3 DEG C, reacts 1h, just can realize being firmly combined between destruction cellulose and lignin
Molecular structure.Because must be destroyed to the structure of Bagasse-cellulose first, bagasse is set to be in " loose " shape
State, to be for further processing.Under more high pressure, and temperature pretreatment condition less high, steam-energy and bagasse
Surface is fully contacted, so as to destroy the molecular structure being firmly combined between cellulose and lignin.
Although being pre-processed by the first step, the structure of bagasse is presented a certain degree of " loose " state, bagasse
Again without sufficiently being degraded, therefore second step uses Microwave Pretreatment.At 120 DEG C, by what is pre-processed through the first step
Bagasse is processed under microwave, may be such that the bagasse in " loose " state is further decomposed, but locate in advance through microwave
After reason, the materials such as some bio oils, diesel oil may be produced, be washed with 90% ethanol, the oily substance of generation can be eluted, just
Grown in the decomposition of next step microbial.
By after first two steps pretreatment, bagasse can carry out the decomposition of the 3rd step, i.e. microorganism and decompose.Microorganism is mainly sharp
With the enzyme system decomposition of cellulose that itself is produced, in the case of cleavage, cellulose is further decomposed.Available point
Solving bacterium isAspergillus niger ATCC 1015、Trichoderma reesei The Hes of ATCC 26921Penicillium janthinellum One kind in ATCC 44750, whereinTrichoderma reesei The discomposing effect of ATCC 26921 is most
Good, this is probably because the bacterium is more biased towards utilizing the decomposition of bagasse.
But several steps essentially consist in decomposability cellulose and hemicellulose above, but some lignin may not obtained also
Sufficiently decompose, therefore, the 4th step is soaked using ammoniacal liquor, the purpose of ammoniacal liquor immersion be make the cellulose that has been destroyed and
Lignin is separated, and has only separated lignin, and the enzymolysis in next step can just make cellulose obtain further decomposing again.
In ammoniacal liquor immersion, soak time, pressure and temperature are more important, by suitable setting, can improve the decomposition effect of bagasse
Rate, and reduce the generation of inhibitor.More than after a few step pretreatments, final step is using enzymolysis.
The order of above-mentioned pre-treatment step is fixed.Processed by above-mentioned steps, can not only obtain product sugar higher
All main inhibitor contents are less than the concentration for influenceing fermentation bacteria growing in rate, and hydrolyzate, or as little as inspection is not measured.
The complexity of decomposition is affected due to the size of bagasse, it is however generally that, smaller bagasse is more easy to decompose, but
In view of Energy Consumption Factors, the size of bagasse is controlled below 860 μm, not only discomposing effect is good, and energy consumption is relatively low.
Liquid hot-water process, without adding acid reagent, will not cause corrosion, the hydrolysis of generation compared to diluted a cid method to reactor
Residue in liquid is also less, is a kind of more gentle preprocess method.However, its sugarcane residue rate of recovery is relatively low, it is single
The purpose for effectively decomposing bagasse is unable to reach when solely using.
After liquid hot-water process is processed, microwave is recycled to be processed, the yield of raising reduced sugar that can be by a relatively large margin,
And shorten reaction time and reducing energy consumption.When carrying out microwave treatment, microwave can accelerate the destruction to crystal structure, make microorganism more
Easily bagasse is further decomposed.After decomposer is decomposed, 10% or so bagasse resolution ratio can be again improved.
Preferably, when bagasse is decomposed using decomposer, utilizeTrichoderma reesei ATCC 26921Can be real
Existing more preferable decomposition efficiency.
When carrying out ammonia treatment, because ammonia is the good solvent of lignin, lignin and lignocellulosic can be separated,
So as to decomposition efficiency of the subsequent fiber element enzyme to bagasse can be improved.
By after above-mentioned treatment, recycling enzymolysis processing, the hydrolysis bagasse of maximizing.
Beneficial effects of the present invention：
1st, can effectively hydrolyzing bagasse, percent hydrolysis can reach 74.41%；
2nd, the glucose of high concentration is can obtain, concentration of glucose can reach 15.21 ± 1.65g/L；
3rd, pretreatment condition is gentle, and energy consumption is low, not etching apparatus；
4th, without following purification steps, microbial fermentation low production cost；
5th, major inhibitors furtural and hydroxymethyl furfural content are low, and the growth of fermentative microorganism is not influenceed, while base
This does not produce other inhibitor such as glucuronic acid, p-Coumaric Acid, syringic acid and forulic acid.
Brief description of the drawings
1st, Fig. 1 show the schematic diagram of preprocess method step of the present invention；
2nd, it is using bagasse hydrolysate shown in Fig. 2（Filtrate）During as fermentation substrate, tunning and fermentation time
Graph of a relation；Wherein HAN is expressed as：In step 4）UsingAspergillus niger ATCC 1015 is incited somebody to action as zymophyte
The filtrate I, filtrate II, filtrate III, filtrate IV and filtrate V merge, as SHF fermentations；HTR is expressed as：In step 4）Adopt
WithTrichoderma reesei ATCC 26921 as zymophyte, and by the filtrate I, filtrate II, filtrate III, filtrate IV
Merge with filtrate V, as SHF fermentations；HPJ is expressed as：In step 4）UsingPenicillium janthinellum
ATCC 44750 merges the filtrate I, filtrate II, filtrate III, filtrate IV and filtrate V as zymophyte, as SHF hairs
Ferment；HANNY is expressed as：In step 4）UsingAspergillus niger ATCC 1015 as zymophyte, and by filtrate I, filter
Liquid II, filtrate III, filtrate IV and solid residue IV merge, as SSF fermentations；HTRNY is expressed as：In step 4）UsingTrichoderma reesei ATCC 26921 as zymophyte, and by filtrate I, filtrate II, filtrate III, filtrate IV and solid
Residue IV merges, as SSF fermentations；HPYNY is expressed as：In step 4）UsingPenicillium janthinellum
Filtrate I, filtrate II, filtrate III, filtrate IV and solid residue IV are merged and rotated dense by ATCC 44750 as zymophyte
Contracting, as SSF fermentations；
3rd, it is using bagasse hydrolysate shown in Fig. 3（Filtrate）During as fermentation substrate, in SSF and SHF fermentation process
The variation diagram of middle pH；Wherein, the implication of HAN, HTR, HPJ, HANNY, HTRNY, HPJNY with it is consistent in Fig. 2；A sends out for SSF
Ferment；B ferments for SHF；
4th, shown in Fig. 4, in being SSF and SHF fermentation process, to the conversion service condition of different sugar composition；Wherein a, b, c are
SHF ferment, b, d, f be SSF fermentation, the implication of HAN, HTR, HPJ, HANNY, HTRNY, HPJNY with it is consistent in Fig. 2；
5th, it is the total residue carbohydrate produced by different fermentations substrate in fermentation termination shown in Fig. 5；Wherein,
The implication of HAN, HTR, HPJ, HANNY, HTRNY, HPJNY with it is consistent in Fig. 2；
6th, shown in Fig. 6, when being by the use of pure glucose as fermentation substrate, during the fermentation, the variation diagram of different parameters；
7th, shown in Fig. 7, when being by the use of sugar composite as fermentation substrate, the variation diagram of different products, sugar and pH.
Specific embodiment
The present invention is specifically described below by embodiment, it is necessary to it is pointed out here that be that following examples are use
It is further detailed in the present invention, it is impossible to be interpreted as limiting the scope of the invention, the field is skilled in technique
Personnel still fall within protection scope of the present invention according to some nonessential modifications and adaptations that foregoing invention content is made.
Embodiment 1：
1）It is 5-10cm that 500g sugarcanes are carried out into segment to length, is dried 2 days at 65 ± 2 DEG C, is then crushed, mistake
20 mesh sieves, obtain bagasse powder of the particle diameter less than 830 μm；
2）By step 1）Gains preheat 45s, Ran Houjia with steam afterwards in 12h is soaked under normal temperature in the water of 2L
Enter liquid hot water, at 200 ± 3 DEG C, react 1h, filtered after reaction, collect filtrate I and solid residue I；
3）Set-up procedure 2）The humidity of gained solid residue is 40%, is placed in three neck round bottom, then passes to nitrogen
Gas excludes oxygen, in 12h is placed at 4 DEG C, is placed in reactor reacts 7 min in microwave-heating stove at 120 DEG C afterwards, arranges
Go out hot gas, after temperature drops to room temperature, reaction gains are embathed with 90% ethanol, filtered afterwards, collect filtrate II
With solid residue II；
4）Baking step 3）Gained solid residue, adds nutrient solution to be mixed afterwards, is gone out at 121 DEG C
Bacterium, after sterilizing, the humidity for adjusting mixture is 30%, adds 20mlAspergillus niger The bacterium solutions of ATCC 1015, in
Ferment 10 d at 30 DEG C, is filtered afterwards, collects filtrate III and solid residue III；
The formula of nutrient solution is：30 mg H3BO3、20 mg MnCl2.4H2O、185 mg ZnSO4•7H2O、20 mg
Na2MoO4•2H2O, 280 mg FeSO4•H2O, 200 mg CuSO4, 5 g NH4NO3, 1 g KH2PO4,0.15 g
MgSO4•H2O, 0.11 g CaCl2 and 1 L ddH2O, pH are 7.0；
5）Use ddH2O is to step 4）Gained solid residue is cleaned, and is baked to afterwards, is 1 with solid-liquid mass ratio:
2, the solid residue III after drying is mixed with ammoniacal liquor, under high pressure（0.1MPa）, in 90 DEG C react 30 min, it is laggard
Row filtering, collects filtrate IV and solid residue IV；
6) by step 5）Gained solid residue IV soaks 2 h in the distilled water of 900ml, with 1% H2SO4Adjusting pH is
7.0, it is concentrated with Rotary Evaporators, it is baked at 60 DEG C.Then enzymolysis mixture is prepared, every 500 ml should
Enzymolysis mixture is made up of following substances：Solid residue IV, 10g cellulases, 100 ml OPTMASH that 50 g have been dried
BG, 5 mg chloramphenicol and 2.5 mg kanamycins；The pH of adjustment enzymolysis mixture is 5.0, is 55 DEG C in temperature, and shaking speed is
Under 220rpm, 60 h are reacted, filtered afterwards, collect filtrate V and solid residue V.
The solid recovery rate of each step is shown in Table 1.
Embodiment 2
1）It is 5-10cm that 500g sugarcanes are carried out into segment to length, is dried 2 days at 65 ± 2 DEG C, is then crushed, mistake
20 mesh sieves, obtain bagasse powder of the particle diameter less than 830 μm；
2）By step 1）Gains preheat 45s, Ran Houjia with steam afterwards in 12h is soaked under normal temperature in the water of 2L
Enter liquid hot water, at 200 ± 3 DEG C, react 1h, filtered after reaction, collect filtrate I and solid residue I；
3）Set-up procedure 2）The humidity of gained solid residue is 40%, is placed in three neck round bottom, then passes to nitrogen
Gas excludes oxygen, in 8h is placed at 4 DEG C, is placed in reactor reacts 7 min in microwave-heating stove at 120 DEG C afterwards, arranges
Go out hot gas, after temperature drops to room temperature, reaction gains are embathed with 90% ethanol, filtered afterwards, collect filtrate II
With solid residue II；
4）Baking step 3）Gained solid residue, adds nutrient solution to be mixed afterwards, is gone out at 121 DEG C
Bacterium, after sterilizing, the humidity for adjusting mixture is 30%, adds 20mlAspergillus niger The bacterium solutions of ATCC 1015, in
Ferment 10 d at 30 DEG C, is filtered afterwards, collects filtrate III and solid residue III；
The formula of nutrient solution is：30 mg H3BO3、20 mg MnCl2.4H2O、185 mg ZnSO4•7H2O、20 mg
Na2MoO4•2H2O, 280 mg FeSO4•H2O, 200 mg CuSO4, 5 g NH4NO3, 1 g KH2PO4,0.15 g
MgSO4•H2O, 0.11 g CaCl2 and 1 L ddH2O, pH are 7.0；
5）Use ddH2O is to step 4）Gained solid residue is cleaned, and is baked to afterwards, is 1 with solid-liquid mass ratio:
2, the solid residue III after drying is mixed with ammoniacal liquor, under high pressure（0.1MPa）, in 90 DEG C react 30 min, it is laggard
Row filtering, collects filtrate IV and solid residue IV；
6) by step 5）Gained solid residue IV soaks 2 h in the distilled water of 900ml, with 1% H2SO4Adjusting pH is
7.0, it is concentrated with Rotary Evaporators, it is baked at 60 DEG C.Then enzymolysis mixture is prepared, every 500 ml should
Enzymolysis mixture is made up of following substances：Solid residue IV, 10g cellulases, 100 ml OPTMASH that 50 g have been dried
BG, 5 mg chloramphenicol and 2.5 mg kanamycins；The pH of adjustment enzymolysis mixture is 5.0, is 55 DEG C in temperature, and shaking speed is
Under 220rpm, 60 h are reacted, filtered afterwards, collect filtrate V and solid residue V.
The solid recovery rate of each step is shown in Table 2.
Embodiment 3
1）It is 5-10cm that 500g sugarcanes are carried out into segment to length, is dried 2 days at 65 ± 2 DEG C, is then crushed, mistake
20 mesh sieves, obtain bagasse powder of the particle diameter less than 830 μm；
2）By step 1）Gains preheat 45s, Ran Houjia with steam afterwards in 12h is soaked under normal temperature in the water of 2L
Enter liquid hot water, at 200 ± 3 DEG C, react 1h, filtered after reaction, collect filtrate I and solid residue I；
3）Set-up procedure 2）The humidity of gained solid residue is 40%, is placed in three neck round bottom, then passes to nitrogen
Gas excludes oxygen, in 10h is placed at 4 DEG C, is placed in reactor reacts 7 min in microwave-heating stove at 120 DEG C afterwards, arranges
Go out hot gas, after temperature drops to room temperature, reaction gains are embathed with 90% ethanol, filtered afterwards, collect filtrate II
With solid residue II；
4）Baking step 3）Gained solid residue, adds nutrient solution to be mixed afterwards, is gone out at 121 DEG C
Bacterium, after sterilizing, the humidity for adjusting mixture is 30%, adds 20mlPenicillium janthinellum ATCC
44750 bacterium solutions, in 10 d that fermented at 30 DEG C, are filtered afterwards, collect filtrate III and solid residue III；
The formula of nutrient solution is：30 mg H3BO3、20 mg MnCl2.4H2O、185 mg ZnSO4•7H2O、20 mg
Na2MoO4•2H2O, 280 mg FeSO4•H2O, 200 mg CuSO4, 5 g NH4NO3, 1 g KH2PO4,0.15 g
MgSO4•H2O, 0.11 g CaCl2 and 1 L ddH2O, pH are 7.0；
5）Use ddH2O is to step 4）Gained solid residue is cleaned, and is baked to afterwards, is 1 with solid-liquid mass ratio:
2, the solid residue III after drying is mixed with ammoniacal liquor, under high pressure（0.1MPa）, in 90 DEG C react 30 min, it is laggard
Row filtering, collects filtrate IV and solid residue IV；
6) by step 5）Gained solid residue IV soaks 2 h in the distilled water of 900ml, with 1% H2SO4Adjusting pH is
7.0, it is concentrated with Rotary Evaporators, it is baked at 60 DEG C.Then enzymolysis mixture is prepared, every 500 ml should
Enzymolysis mixture is made up of following substances：Solid residue IV, 10g cellulases, 100 ml OPTMASH that 50 g have been dried
BG, 5 mg chloramphenicol and 2.5 mg kanamycins；The pH of adjustment enzymolysis mixture is 5.0, is 55 DEG C in temperature, and shaking speed is
Under 220rpm, 60 h are reacted, filtered afterwards, collect filtrate V and solid residue V.
The solid recovery rate of each step is shown in Table 3.
Embodiment 4
1）Substep diastatic fermentation method：Respectively by the filtrate I in the middle of embodiment 1, embodiment 2 and embodiment 3, filtrate II, filter
Liquid III, filtrate IV and filtrate V merge and concentrate.In each embodiment, the sugared composition of the filtrate after merging is as shown in table 4.
Filtrate after 150ml is merged is placed in the anaerobic of 250ml specifications bottle, and is sealed with butyl rubber plug；Add 0.75g
Yeast extract and 1.5g peptones, pH to 6.5 is adjusted with 1% NaOH, after sterilizing 20 min in 115 DEG C, is cooled to normal temperature；
The mixed solution after 0.5 ml sterilizings is added, the mixed solution contains 50g/L KH2PO4、50g/L K2HPO4、220g/L
CH3COONH4,0.1g/L p-aminobenzoic acid, 0.1g/L vitamin Bs, 0.001g/L biotins, 20 g/L MgSO4 •7H2O、
1 g/L MnSO4 •H2O、1 g/L FeSO4 •7H2O and 1 g/L NaCl；Then ferment to OD600=1.5.
2）Simultaneous saccharification and fermentation method：Respectively by the filtrate I in the middle of embodiment 1, embodiment 2 and embodiment 3, filtrate II, filter
Liquid III and filtrate IV merge, and adjust pH to 7.0.In each embodiment, the composition of the filtrate after merging is as shown in table 5.
Fermentation step is carried out simultaneously with enzymolysis, and added trace element and fermentation condition are consistent with distribution diastatic fermentation method.Hair
Ferment temperature is 37 DEG C, and fermentation time is 96h, during the fermentation, every a period of time, is carried out through rubber stopper with asepsis injector
Sampling, samples taken is centrifuged 20min at 4 DEG C under 8000rpm, then carry out the detection of sugar, organic acid, pH and ABE production.
Embodiment 5
The detection of inhibitor content：In the liquid chromatographs of DIONEX UltiMate 3000, using Aminex HPX-87H
Chromatographic column is detected to butyric acid, acetic acid, glucuronic acid, p-Coumaric Acid, syringic acid and ferulaic acid content；Furtural and 5- hydroxyls
The measure of methyl furfural refers to " Determination of 5-Hydroxymethylfurfural in Vinegar
Samples by HPLC”（Theobald A, Müller A, Anklam E. J. Agric. Food. Chem. 1998,
46(5):1850-1854.）In method.
After all pre-treatment steps, each inhibitor content is shown in Table 6 in embodiment 1-3.
Comparative example
Under fermentation condition same as Example 4, using pure glucose or sugar composite（10g/L glucose, 42g/L xyloses,
3.5g/L arabinoses, 3.5g/L cellobioses, 3g/L galactolipins and 1.5g/L mannoses）Substitute the hydrolysate of pretreatment
（Filtrate）.
Embodiment 6
Tunning is detected：Alcohols material（Acetone, butanol, ethanol）Detected using gas chromatograph；Total solid residue
The detection of carbohydrate refers to " Carbohydrate analysis by a phenol-sulfuric acid method
in microplate format”（Masuko T, Minami A, Iwasaki N, Majima T, Nishimura S-I,
Lee YC: Anal. Biochem. 2005, 339(1):69-72.）In method；Content of cellulose utilizes spectrophotometer
Detected；The detection of content of lignin refers to " Determination of lignin in herbaceous plants by
an improved acetyl bromide procedure”（Iiyama K, Wallis AFA J. Sci. Food
Agric. 1990, 51(2):145-161.）In method；Content of ashes is examined after being dried in 500 DEG C in muffle kiln
Survey.
In fermentation process, each parameter index of tunning is shown in Table 7.
Claims (2)
1. the preprocess method of butanol is produced in a kind of utilization bagasse fermentation, it is characterised in that：Methods described comprises the following steps：
1）Mechanical crushing：It is 5-10cm that segment to length is carried out to sugarcane, is dried 2 days at 65 ± 2 DEG C, is then crushed,
20 mesh sieves are crossed, bagasse powder of the particle diameter less than 830 μm is obtained；
2）Liquid hot water treatment：It is 250g/L by solid-liquid mass volume ratio, by step 1）Gains are in the immersion in water under normal temperature
12h, preheats 45s with steam afterwards, is subsequently adding liquid hot water, at 200 ± 3 DEG C, reacts 1h, is filtered after reaction,
Collect filtrate I and solid residue I；
3）Microwave treatment：Set-up procedure 2）The humidity of gained solid residue I is 40%, is placed in reactor, then passes to nitrogen
Gas excludes oxygen, in 8-12h is placed at 4 DEG C, reactor is placed in microwave-heating stove afterwards, and 7 are reacted at 120 DEG C
Min, then reaction gains are cooled to normal temperature, reaction gains are embathed with 90% ethanol, filtered afterwards, collect
Filtrate II and solid residue II；
4）Microorganism is decomposed：Baking step 3）Gained solid residue II, adds nutrient solution to be mixed, then at 121 DEG C afterwards
Under sterilized, after sterilizing, adjust its humidity for 30%, by percentage to the quality, 10% decomposer bacterium solution is added, in 30 DEG C
10 d of lower fermentation, are filtered afterwards, collect filtrate III and solid residue III；
The formula of the nutrient solution is：30 mg H3BO3、20 mg MnCl2.4H2O、185 mg ZnSO4•7H2O、20 mg
Na2MoO4•2H2O、280 mg FeSO4•H2O、200 mg CuSO4、5 g NH4NO3、1 g KH2PO4、0.15 g MgSO4•
H2O、0.11 g CaCl2With 1 L ddH2O, pH are 7.0；
The decomposer isAspergillus niger ATCC 1015、Trichoderma reesei The Hes of ATCC 26921Penicillium janthinellum One kind in ATCC 44750；
5）Ammonia treatment：Use ddH2O is to step 4）Gained solid residue III is cleaned, and is baked to afterwards, with solid-liquid mass ratio
It is 1:2, the solid residue III after drying is mixed with ammoniacal liquor, under 0.1MPa pressure, 30 min are reacted in 90 DEG C, afterwards
Filtered, collected filtrate IV and solid residue IV；
6）Enzymolysis processing：The solid residue IV is soaked into 2 h in water, regulation pH is 7.0, is baked to after concentration, so
Enzymolysis mixture is prepared afterwards, and every 500 ml enzymolysis mixtures are made up of following substances：The solid residues that 50 g have been dried
Thing IV, 10g cellulases, 100 ml OPTMASH BG, 5 mg chloramphenicol and 2.5 mg kanamycins；Adjustment enzymolysis mixture
PH be 5.0, temperature be 55 DEG C, shaking speed be 220rpm under, react 60 h, filtered afterwards, collect the He of filtrate V
Solid residue V；
7）Filtrate I, filtrate II, filtrate III, filtrate IV and solid residue IV are merged, SSF fermentations are remained, or by the filtrate
Ith, filtrate II, filtrate III, filtrate IV and filtrate V merge and concentrated by rotary evaporation, remain SHF fermentations.
2. method according to claim 1, it is characterised in that：In methods described, step 4）In decomposer used beTrichoderma reesei ATCC 26921。
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201410744063.8A CN104357491B (en) | 2014-12-09 | 2014-12-09 | The preprocess method of butanol is produced in a kind of utilization bagasse fermentation |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201410744063.8A CN104357491B (en) | 2014-12-09 | 2014-12-09 | The preprocess method of butanol is produced in a kind of utilization bagasse fermentation |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN104357491A CN104357491A (en) | 2015-02-18 |
| CN104357491B true CN104357491B (en) | 2017-07-07 |
Family
ID=52524799
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201410744063.8A Active CN104357491B (en) | 2014-12-09 | 2014-12-09 | The preprocess method of butanol is produced in a kind of utilization bagasse fermentation |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN104357491B (en) |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106906258A (en) * | 2017-03-01 | 2017-06-30 | 贵州大学 | A kind of method for producing 5 hydroxymethylfurfurals |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103993053A (en) * | 2014-05-07 | 2014-08-20 | 中国科学院广州能源研究所 | Method for pretreatment of biomass by water-ammonia coupling |
| WO2014144588A1 (en) * | 2013-03-15 | 2014-09-18 | Suganit Systems, Inc. | Alkaline treatment of lignocellulosic biomass |
-
2014
- 2014-12-09 CN CN201410744063.8A patent/CN104357491B/en active Active
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2014144588A1 (en) * | 2013-03-15 | 2014-09-18 | Suganit Systems, Inc. | Alkaline treatment of lignocellulosic biomass |
| CN103993053A (en) * | 2014-05-07 | 2014-08-20 | 中国科学院广州能源研究所 | Method for pretreatment of biomass by water-ammonia coupling |
Non-Patent Citations (1)
| Title |
|---|
| "微波前处理对水稻秸秆酶解性能的影响";沈洁等;《安徽农业科学》;20140910;第42卷(第26期);摘要，材料与方法 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN104357491A (en) | 2015-02-18 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Khaleghian et al. | Ethanol production from rice straw by sodium carbonate pretreatment and Mucor hiemalis fermentation | |
| US10513714B2 (en) | Lignocellulosic conversion process comprising sulfur dioxide and/or sulfurous acid pretreatment | |
| EP2625279B1 (en) | Enzymatic hydrolysis of lignocellulosic material in the presence of sulfur oxyanions and/or sulfhydryl compounds | |
| EP2449092B1 (en) | Biomass hydrolysis process | |
| Láinez et al. | Bioethanol production from enzymatic hydrolysates of Agave salmiana leaves comparing S. cerevisiae and K. marxianus | |
| CA2833194C (en) | Methods for improvement of enzymatic hydrolysis of lignocellulosic material | |
| Favaro et al. | Processing wheat bran into ethanol using mild treatments and highly fermentative yeasts | |
| Montipó et al. | Integrated production of second generation ethanol and lactic acid from steam-exploded elephant grass | |
| Zhu et al. | Conversion of aqueous ammonia-treated corn stover to lactic acid by simultaneous saccharification and cofermentation | |
| JP2010510800A (en) | Process for fermentative production of ethanol from solid lignocellulosic material comprising the step of treating the solid lignocellulosic material with an alkaline solution to remove lignin | |
| Lienqueo et al. | Second generation bioethanol from Eucalyptus globulus Labill and Nothofagus pumilio: ionic liquid pretreatment boosts the yields | |
| US20230121068A1 (en) | Processes for co-producing xylitol with ethanol or other fermentation products | |
| WO2010077170A2 (en) | Process and system for production of organic solvents | |
| Molaverdi et al. | Efficient ethanol production from rice straw through cellulose restructuring and high solids loading fermentation by Mucor indicus | |
| Vincent et al. | Simultaneous saccharification and fermentation of ground corn stover for the production of fuel ethanol using Phanerochaete chrysosporium, Gloeophyllum trabeum, Saccharomyces cerevisiae, and Escherichia coli K011 | |
| JP5701632B2 (en) | Sugar-containing composition | |
| CN113614239A (en) | Method for treating lignocellulosic biomass | |
| CN104357491B (en) | The preprocess method of butanol is produced in a kind of utilization bagasse fermentation | |
| JP2014158437A (en) | Saccharified solution of lignocellulosic biomass, and manufacturing and application method thereof | |
| Vincent et al. | Evaluation of potential fungal species for the in situ simultaneous saccharification and fermentation (SSF) of cellulosic material | |
| Seo et al. | Improved bioethanol production using activated carbon-treated acid hydrolysate from Corn Hull in Pachysolen tannophilus | |
| CN105647980B (en) | A kind of method that lignocellulosic enzymatic hydrolysis and fermentation produces butanol | |
| KR101412351B1 (en) | Method for preparing ethanol using ethanol fermentation of whole slurry of pretreated lignocelluloses | |
| JP2015042145A (en) | Method for producing ethanol | |
| JP5999759B2 (en) | Biomass processing method |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |