CN103014094A - Pre-treatment method for improving enzymolysis efficiency of plant fiber raw material - Google Patents
Pre-treatment method for improving enzymolysis efficiency of plant fiber raw material Download PDFInfo
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- CN103014094A CN103014094A CN2012105942359A CN201210594235A CN103014094A CN 103014094 A CN103014094 A CN 103014094A CN 2012105942359 A CN2012105942359 A CN 2012105942359A CN 201210594235 A CN201210594235 A CN 201210594235A CN 103014094 A CN103014094 A CN 103014094A
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Abstract
The invention discloses a pre-treatment method for improving the enzymolysis efficiency of a plant fiber raw material, comprising the following steps of sequentially carrying out fungus degradation treatment and hydrolysis treatment of FeCl3 on lignocellulose-based biomass. According to the method disclosed by the invention, fungus pre-treatment and a physiochemical method are combined to overcome the defect of independently pre-treating funguses; and a pre-treatment manner of combining the fungus pre-treatment and the physiochemical method cooperate with each other, so that the yield of a final product is improved.
Description
Technical field
The present invention relates to a kind of method of using microorganism to prepare compound, particularly a kind of method of using microorganism and the preparation of compound synergy to contain the compound of saccharide residue belongs to technical field of biochemical industry.
Background technology
Aggravation along with the day by day in short supply and global warming trend of petroleum resources has become inexorable trend by production of renewable energy resources fuel.In the face of this challenge, the most feasible selection is exactly to utilize s-generation bio-ethanol technology, does raw material with agriculture and forestry organic waste material and special-purpose energy crop and produces thing ethanol next life.By enzymic hydrolysis and two kinds of operations of fermentation lignocellulose being converted into ethanol is present the most feasible mode, and the turnout of bio-ethanol is large, and low production cost.
But the main component of lignocellulose based biomass is Mierocrystalline cellulose, hemicellulose and lignin.When adopting the hydrolyzing ligno-cellulose with cellulosic enzyme raw material, cellulase must just can make reaction carry out on cellulosic substrate by Contact-sorption, and therefore, Mierocrystalline cellulose is the key factor that determines hydrolysis rate to the accessibility of cellulase.And the existence of lignin has hindered the accessibility of Mierocrystalline cellulose to enzyme, and the multiple component structure of the condition of surface of cellulosic crystalline texture and Woody Biomass, Woody Biomass, lignin are caused lignocellulosic material to be difficult to hydrolysis to cellulosic provide protection and Mierocrystalline cellulose by the factor of the structure such as hemicellulose covering and chemical composition.For thereby the enzymic hydrolysis efficient that improves lignocellulose reaches higher ethanol production, raw material must be through suitable pre-treatment before carrying out the enzymic hydrolysis processing.Effectively pretreatment mode must improve Mierocrystalline cellulose to the accessibility of enzyme, and reducing simultaneously has inhibiting degraded product to follow-up fermenting process.At present, the pretreated method of lignocellulosic material mainly contains Physical, chemical method, physico-chemical processes and biological process.
Traditional Physical comprises mechanical crushing method; High-temperature decomposition; The technology such as high energy electron irradiation, microwave, ultrasonic wave.Chemical method is divided into dilute acid pretreatment method, Alkaline pretreatment method, ozone treatment method, organic solvent treatment process, oxidation treatment method.Physico-chemical processes has steam explosion, ammonia fiber blasting procedure, carbonic acid gas blasting procedure etc., energy consumption was large when but above-mentioned pretreatment process was processed plant fiber material, etching apparatus, production cost is expensive, and follow-up enzymolysis processing there is restraining effect, microorganism is had toxicity, and environmental pollution is serious in addition, non-long-term solution.The contact area of enzyme-to-substrate raw material has improved enzymolysis efficiency when having increased enzymolysis processing after for example mechanical crushing method is pulverized lignocellulose raw material, but reduces to some extent with the impact that the sample smashing fineness increases its his-and-hers watches area change.So lignocellulosic particles tiny to a certain extent after, continue to pulverize and can only limitedly improve enzymolysis efficiency, and processing cost increases relatively.And for example, although the ultrasonication technology can effectively be destroyed the hydrogen bond in the cellulosic molecule, reduce its crystallization degree and regularity, the fiber of crossing through ultrasonication can increase the accessibility to cellulase and xylosidase, and is favourable to enzymic hydrolysis; But limited on cellulosic microtexture impact, and the hemicellulose of having degraded, causing the decline of pulp freeness, this point is disadvantageous to enzymic hydrolysis.
Biological process is to utilize microorganism (such as fungi, actinomycetes, bacterium etc.) lignocellulose degradation based biomass, the biological process action condition is gentle, energy consumption is low, need not to add chemical reagent and without the output of waste, can improve significantly enzymic hydrolysis efficient, have unique advantage, but the treatment time of biological process preprocessing lignocellulose raw material is long, production cycle is long, and processing efficiency is low, has consumed carbohydrate simultaneously in lignin degrading.
Summary of the invention
Primary and foremost purpose of the present invention is to provide a kind of pretreatment process that improves the plant fiber material enzymolysis efficiency for the problem that above-mentioned prior art exists, the inventive method combines bioremediation and physico-chemical process can solve biology or the independent pretreated defective of physico-chemical process, two kinds of successively combinations of pretreatment process, pre-treatment to plant fiber material has synergistic effect, improve Mierocrystalline cellulose to the accessibility of enzyme, reduce the restraining effect effect in the follow-up enzymolysis processing process of biomass, improved the yield of biomass degradation end product reducing sugar.
In order to achieve the above object, one aspect of the present invention provides a kind of pretreatment process that improves the plant fiber material enzymolysis efficiency, comprises that plant fiber material is carried out fungus degrading successively to be processed and FeCl
3Chemical hydrolysis process.
Wherein, described fungus degrading process be with plant fiber material with after the fungi inoculation liquid is mixed, be under 28 ± 3 ℃ in temperature, cultivated 20-40 days.
Particularly, described fungi is selected brown rot fungus (Fomitopsis palustris C7615).
Particularly, described plant fiber material is prepared from as follows: at first plant fiber material is pulverized; Then the water ratio of regulating plant fibrous material is to 70-80%; Then plant fiber material is carried out sterilising treatment, and get final product.
Especially, the temperature of described sterilising treatment is 121 ± 1 ℃; Sterilization time is 15-25min.
Wherein, the granularity of the plant fiber material after the pulverizing is the 20-80 order.
Particularly, also comprise plant fiber material is dried to water ratio≤10% after, carry out again described pulverizing.
Wherein, the volume of fungi inoculation suspension is 10-20:100 with the ratio of the weight of plant fiber material, be preferably 15:100, namely inoculate the described brown rot fungus inoculation of 10-20L suspension in every 100g pre-treatment material in the described brown rot fungus inoculation suspension of inoculation 10-20ml or the every 100kg pre-treatment material.
Particularly, described fungi inoculation liquid is prepared from accordance with the following steps: at first fungi is inoculated in the amplification culture base, under dark condition, in 28 ± 1 ℃, rotating speed is to cultivate 3-7 days under the condition of 100-150rpm, then adds sterile distilled water, carry out homogenized, and get final product.
Especially, the rotating speed in the described amplification culture process is preferably 150rpm; Incubation time is preferably 5 days; The rotating speed of described homogenized is 4000-5000rpm; The homogenized time is 30-50s.
Wherein, described amplification culture based component: 20g glucose, 5g yeast soak powder, 1gKH
2PO
4, 0.5gMgSO
4, the 0.01g vitamins B
1, it is 1000ml that deionized water is regulated cumulative volume.Cooling is stand-by after the sterilization.
Particularly, the ratio of the volume of the amplification culture base that the volume of the sterile distilled water that adds uses during with inoculation is 0.7-1.2:1, be preferably 1:1, the volume of the amplification culture base that namely uses when inoculation is 100ml, then add the 100ml sterile distilled water, if the volume of the amplification culture base that uses during inoculation is 1L, then add the 1L sterile distilled water.
Wherein, the composition of the described amplification culture base of every 1000ml consists of: 20g glucose, 5g yeast soak powder, 1gKH
2PO
4, 0.5gMgSO
4, the 0.01g vitamins B
1, it is 1000ml that deionized water is regulated cumulative volume.Cooling is stand-by after the sterilization.
Particularly, also comprise the plant fiber material after the fungus degrading processing is carried out drying treatment, wherein drying temperature is 60-70 ℃, and be 1-2 days time of drying.
Especially, the fibrous material water ratio after dried fungi is processed is 5%-7%.
Wherein, described FeCl
3Chemical hydrolysis process and to comprise following in sequence step:
A) with plant fiber material and FeCl after the fungus degrading processing
3Solution mixes, and makes FeCl
3The hydrolysis mixed solution;
B) heating FeCl
3The hydrolysis mixed solution is that to carry out chemical hydrolysis reaction under 140-180 ℃ be 20-30min keeping temperature;
C) to the processing of lowering the temperature of the mixture behind the hydrolysis reaction, until the temperature of hydrolysis reaction mixture reaches 15-35 ℃.
The weight of the plant fiber material particularly, steps A) behind the fungus degrading and FeCl
3The ratio of the volume of solution is 7.5-15:100, is preferably 10:100; Described FeCl
3The concentration of solution is 0.05-0.2mol/L, is preferably 0.1mol/L; Step B) temperature of chemical hydrolysis described in is preferably 160-180 ℃, more preferably 180 ℃; Hydrolysis time is preferably 25-30min, more preferably 30min.
Particularly, comprise that also the plant fiber material to hydrolysis treatment carries out drying treatment, drying temperature is 25-40 ℃, is preferably 35 ℃; Be 1-3 days time of drying, is preferably 2 days.
Especially, to process the plant fiber material water ratio be 5%-7% for dried chemical hydrolysis.
Particularly, comprise that also the plant fiber material that chemical hydrolysis is processed filters, washing, until the purge flow fluid is neutral, namely the pH value of effluent liquid is to carry out described drying treatment after 7 again.
Pretreatment process of the present invention has following advantage:
1, the inventive method combines bioremediation and physico-chemical process and can solve biology or the independent pretreated defective of physico-chemical process, two kinds of successively combinations of pretreatment process, pre-treatment to the lignocellulose based biomass has synergistic effect, improve Mierocrystalline cellulose to the accessibility of enzyme, reduce the restraining effect effect in the follow-up enzymolysis processing process of biomass, improved the yield of biomass degradation end product reducing sugar.
2, the plant fiber material among the present invention passes through brown rot fungus and FeCl successively
3After the pre-treatment, the enzymic hydrolysis efficient of plant fiber material improves, and carbohydrate transforms fully in the timber, and the yield of reducing sugar significantly improves, the yield of reducing sugar reaches more than 95.36%, even the carbohydrate in the plant fiber material all can be converted into reducing sugar.
3, the inventive method processing step is simple, only need through two step simple process, processing ease, simple to equipment requirements, temperature of reaction is low, and reaction conditions is gentle, the pre-treatment less energy consumption, the pollutent that produces is few, can greatly improve the enzymic hydrolysis efficient of lignocellulose, is extremely feasible Pretreatment Technologies of Lignocellulose during a kind of extension in future is produced.
4, the pre-treatment of brown rot fungus fungi utilizes can degrade part hemicellulose and make the action principle of xylogen generation depolymerization of brown rot fungus in the inventive method, the Mierocrystalline cellulose in the plant fiber material is increased greatly to the accessibility of enzyme, thereby improved enzymic hydrolysis efficient.
Description of drawings
Fig. 1 is that reference examples 1 and reference examples 2-4 are through FeCl
3The X-ray diffractogram of degradation treatment Cortex Populi Tomentosae powder, wherein, A1 is that reference examples 1 Cortex Populi Tomentosae powder, A2 are that reference examples 2 Cortex Populi Tomentosae powder, A3 are that reference examples 3 Cortex Populi Tomentosae powder, A4 are reference examples 4 Cortex Populi Tomentosae powder;
Fig. 2 is that embodiment 1-3 is successively through fungi, FeCl
3Degradation treatment Cortex Populi Tomentosae powder, reference examples 5 be the X-ray diffractogram of fungus degrading processing Cortex Populi Tomentosae powder only, and wherein B1 is that embodiment 1 Cortex Populi Tomentosae powder, B2 are that embodiment 2 Cortex Populi Tomentosae powder, B3 are that embodiment 3 Cortex Populi Tomentosae powder, B4 are reference examples 5 Cortex Populi Tomentosae powder;
Fig. 3 is the scanning electron microscope (SEM) photograph of Cortex Populi Tomentosae powder, and wherein, A is that fungus degrading is processed the sample scanning electron microscope (SEM) photograph among the embodiment 3; B is reference examples 1 unprocessed Sample Scan Electronic Speculum figure; C is reference examples 4 Sample Scan Electronic Speculum figure; D is through the collaborative FeCl of fungi among the embodiment 3
3Sample Scan Electronic Speculum figure after the solution-treated.
Embodiment
Further describe the present invention below in conjunction with specific embodiment, advantage and disadvantage of the present invention will be more clear along with description.But these embodiment only are exemplary, scope of the present invention are not consisted of any restriction.It will be understood by those skilled in the art that lower without departing from the spirit and scope of the present invention and can make amendment or replace the details of technical solution of the present invention and form, but these modifications and replacing all fall within the scope of protection of the present invention.
The bacterial strain that the present invention uses is brown rot fungus Fomitopsis palustris C7615, picks up from Chinese Guangdong in 2009 and economizes the Che Baling wilderness area, and gathering the people is that Beijing Forestry University's Cui's treasured is triumphant.Strain name is worn by Beijing Forestry University and is kindly helped secure the success of professor's evaluation.Bacterial strain refrigerates the institute of microbiology in Beijing Forestry University under 4 ℃ of conditions after separation and purification.
The Agar Plating composition of brown rot fungus: 20g Fructus Hordei Germinatus soaks powder, 10g glucose, 3g KH
2PO
4, 18g agar, it is 1000ml that deionized water is regulated cumulative volume.Cooling is stand-by after the sterilization.
The amplification culture based component of brown rot fungus: 20g glucose, 5g yeast soak powder, 1gKH
2PO
4, 0.5gMgSO
4, the 0.01g vitamins B
1, it is 1000ml that deionized water is regulated cumulative volume.Cooling is stand-by after the sterilization.
The separation of brown rot fungus: the mycelia that selects survival in the fresh sporophore of this bacterium, be inoculated on the Agar Plating, place 28 ℃ incubator to leave standstill cultivation, observe the mycelial growth situation: if mycelia pollutes, then continue to select dull and stereotyped clean mycelia and again be inoculated in the new flat board, until mycelial growth is good, pollution-free.
Brown rot fungus Fomitopsis palustris morphological specificity: sporophore: the annual or lasting growth of basidioma, calm to calm warp, semicircle, only or imbricate is arranged; Single cap is flat to trilateral, and 10 * 5 * 3 centimetres, upper surface white is to cream-colored, along with the increase at age or become faint yellowly after doing, unconspicuous endless belt and shallow ridges is arranged, and fine hair is arranged or without hair, gradual change is smooth; Edge and cap are homochromy.Vent surface white is to cream-colored, along with the age increase or do after become light yellowly, circular to polygon, most of 2 ~ 4 every millimeter, what have large reaches 1 every millimeter, thin-walled, complete or indentation slightly.Bacterial context is cream-colored, and without ring grain, the stiff fibre shape is to suberin, and thick 2 centimetres, tube and bacterial context are homochromy, thick 1 centimetre.Give out the stench as the rubbish when fresh.
Mycelial structure: mycelia system two is type, and the bacterial context generative hyphae is as thin as heavy wall, and is transparent, has clamp connexion, few branch, and 2.5 ~ 4 microns of diameters, the bacterial context skeletal hyphae is transparent, and heavy wall is without simply separating few branch, 3 ~ 6 microns of diameters; The tube mycelia is similar to the bacterial context mycelia.
Cystidium and load: cystidium and other sterile thalamium structure without.The load club-like, 4 load stalks of tool, 24 ~ 28 * 6 ~ 7 microns, base portion has clamp connexion.
Spore: sporidium is cylindrical, and the frequent hettocyrtosis of base portion is transparent, smooth, without having a liking for blue reaction, 6 ~ 8 * 2.5 ~ 3 microns.
Decaying type: deciduous tree and softwood tree dry wood brown rot.
As example, other lignin-base biomass all are applicable to the present invention to the lignin-base biomass of using in the embodiment of the invention, such as poplar, Eucalyptus, birch, pine, wheat straw, straw, maize straw etc. with Cortex Populi Tomentosae timber; The cellulase that the present invention uses, β-Portugal's glycosides carbohydrase are available from Sigma aldrich company.
Embodiment 1
1, preparation fungi inoculation liquid
Be inoculated on the dull and stereotyped activation medium refrigerating in the brown rot fungus under 4 ℃, carried out activation culture 4-7 days, then the brown rot fungus of getting after the activation is inoculated in the amplification culture base of 100mL in right amount, under 28 ℃ of conditions, add sterile distilled water 100mL in the rotating speed cultivation with 150rpm on the shaking table after 5 days, then stirred 30 seconds with the speed of refiner with 5000rpm, make brown rot fungus inoculation suspension.
The fungi inoculation liquid carries out that culture temperature is 28 ± 1 ℃ in the amplification culture process among the present invention, and the rotating speed of shaking table is that 100-150rpm, incubation time are 3-7 days, then carries out the bacteria suspension that homogenized obtains and all is applicable to the present invention.
2, the pulverization process of raw material
With air-dry to water ratio be 10% Cortex Populi Tomentosae (Populus tomentosa) wood powder to be broken into particle diameter be 20-80 purpose Cortex Populi Tomentosae dry powder, then in the Cortex Populi Tomentosae dry powder, add entry, make water ratio and be 70% pre-treatment material.
3, fungus degrading is processed
Be that 70% Cortex Populi Tomentosae pre-treatment material places Autoclave with water ratio, heating is under 121 ℃ of conditions keeping temperature, and then sterilization 20min is cooled to room temperature (25 ℃), makes the sterilized plant fibrous material;
Brown rot fungus is inoculated suspension to be inoculated in the sterilized plant fibrous material, then in dark, temperature is under 28 ℃ the condition, cultivated 30 days, carry out fungi fermentation, degraded, wherein the volume of brown rot fungus inoculation suspension is 15:100 with the ratio of the weight of pre-treatment material, namely inoculates the described brown rot fungus inoculation of 15L suspension in every 100g pre-treatment material in the described brown rot fungus inoculation suspension of inoculation 15ml or the every 100kg pre-treatment material; Then the plant fiber material after the brown rot fungus degradation treatment is filtered, filter residue carries out drying in baking oven, make the fungus degrading fibrous material, and wherein drying temperature is 60 ℃, and be 2 days time of drying, and dried fungus degrading fibrous material water ratio is 7%.
4, chemical degradation is processed
It is the reactor (the mechanical company limited of the prosperous Sheng in Zhejiang) of 50mL that the fungus degrading fibrous material is placed the cubic capacity by the stainless steel casting, then adds the FeCl that concentration is 0.1mol/L
3Solution, the wherein weight of fungus degrading fibrous material and FeCl
3The ratio of the volume of solution is 10:100;
Reactor is placed the IKA magnetic force well heater (C-MAG HS7) that oil bath is installed, reactor is carried out the oil bath heating, so that the temperature in the reactor remains 140 ℃, fibrous material carries out chemical degradation reaction 30min, then take out reactor, be the tap water sprinkle reactor of (10-30 ℃) with temperature, until the temperature in the reactor drops to room temperature (25 ℃);
Filter the reaction degradation product in the reactor, then repeatedly clean undissolved residue with distilled water, until the purge flow fluid is neutral, namely the pH value of effluent liquid is 7; Then the residue after will cleaning carries out drying and processing under 35 ℃, drying time is 2 days, and the water ratio of the residue after the oven dry is 7%, namely obtains FeCl
3The degradation of fibers raw material.
Measure the collaborative FeCl of fungi according to the standard method in U.S. renewable energy source laboratory
3Mierocrystalline cellulose after processing in the fibrous material, the content of hemicellulose, namely add mass percent concentration in the fibrous material after process and be 72% sulfuric acid, be under 30 ℃ in temperature, behind the hydrolysis 1h, adding deionized water, above-mentioned substance is diluted to concentration is 4% mixed liquor, then mixed liquor being put in the Autoclave, is under 121 ℃ the condition in temperature, heating 1h.Utilize poor heavy method to measure content of lignin.
Utilize high performance liquid phase ion chromatograph (Dai An company) to measure FeCl
3Mierocrystalline cellulose in the degradation of fibers raw material, the content of hemicellulose.High-efficiency anion exchange ion chromatographic column (ICS3000 wears peace, the U.S.) is equipped with pulsed amperometric detector (CAD), AS50 automatic sampler, glycan analysis post (PA-20,4 * 250mm wear peace) and guard column (PA-20,3 * 30mm wear peace).The concrete analysis condition is: adopt the drip washing of 2.75mM NaOH isoconcentration to measure 6 kinds of contents of monosaccharides, flow velocity is 0.4mL/min, and the drip washing time is 40min.Adopt subsequently the drip washing of 1M NaAc isoconcentration to measure 2 kinds of glucuronic acid contents, flow velocity is 0.4mL/min, and the drip washing time is 10min.Adopt at last 137.4mM NaOH to clean chromatographic column 10min.The content of monose and uronic acid is obtained by the typical curve conversion of concentration known.Wherein the monose standard specimen is L-rhamnosyl, L-arabinose, D-Glucose, D-wood sugar, D-MANNOSE and D-semi-lactosi, and the standard specimen of uronic acid is D-Glucose aldehydic acid and D-galacturonic acid.Wherein, the content of sample fiber element is that content with glucose that this sample is surveyed multiply by hydrolysis coefficient 0.9 and obtains; The content of hemicellulose is the summation of wood sugar that sample is surveyed, semi-lactosi and pectinose, and wherein wood sugar and galactose on hydrolysis coefficient are 0.88, and pectinose hydrolysis coefficient is 0.9.Measurement result is as shown in table 1.
Adopting concentration in the embodiment of the invention is the FeCl of 0.1mol/L
3Solution carries out chemical degradation and processes FeCl
3The concentration of solution is except 0.1mol/L, and other concentration are the FeCl of 0.05-0.2mol/L
3Solution also is applicable to the present invention.
5, enzymic hydrolysis is processed
In 10mL sodium acetate buffer (50mM, pH4.8), add the tetracycline of 40 μ L and the cycloheximide solution of 20 μ L, mix, make the enzymic hydrolysis solvent, for subsequent use, wherein the concentration of tetracycline is 10%(m/v), the concentration of cycloheximide solution is 10% (m/v);
Add the 0.2g(dry weight to the enzymic hydrolysis solvent) FeCl
3Preprocessing biomass, cellulase and β-Portugal's glycosides carbohydrase stirs to be placed on and carries out enzymic hydrolysis on the shaking table and process 96h and namely get enzymic hydrolysate, and its cellulase is that the 35FPU/g-substrate (is FeCl
3Preprocessing biomass), β-Portugal's glycosides carbohydrase is the 37.5IU/g-substrate, and the temperature that enzymic hydrolysis is processed is 50 ℃, and rotating speed is 150rpm.
Enzymic hydrolysate is carried out centrifugal treating, get supernatant liquor after centrifugal and carry out reducing sugar content according to DNS method (dinitrosalicylic acid method) and measure, calculate Reducing sugar, calculation formula is as follows:
Wherein, the total amount of Mierocrystalline cellulose and hemicellulose is the summation of Mierocrystalline cellulose and hemicellulose in the Cortex Populi Tomentosae fibrous material after chemical degradation is processed in the sample, records according to the standard method in U.S. renewable energy source laboratory; 0.9 be the hydrolysis coefficient.
Reducing sugar is as shown in table 2.
Embodiment 2
In the pulverization process step of raw material except with water ratio be 7% Cortex Populi Tomentosae wood powder to be broken into granularity be 20-80 purpose powder, adding water, to make water ratio be that all the other are identical with embodiment 1 outside 80% the pre-treatment material;
The fungus degrading treatment step is 20:100 except the ratio of the volume of brown rot fungus inoculation suspension and the weight of pre-treatment material, i.e. the described brown rot fungus inoculation of inoculation 20L suspension in the described brown rot fungus inoculation suspension of inoculation 20ml or the every 100kg pre-treatment material in every 100g pre-treatment material; Culture temperature is 25 ℃, and incubation time is 40 days; Drying temperature is 70 ℃, and be 1 day time of drying, and dried fungus degrading fibrous material water ratio is outside 5%, and all the other are identical with embodiment 1;
The chemical degradation treatment step remains 160 ℃ except the temperature of reaction of reactor, the DeR time is 25min, tap water sprinkle reactor, until the temperature in the reactor drops to room temperature (15 ℃), residue after the cleaning carries out drying and processing under 25 ℃, drying time is 3 days, and the water ratio of the residue after the oven dry is outside 6%, all the other are identical with embodiment 1, measure the collaborative FeCl of fungi according to the standard method in U.S. renewable energy source laboratory
3Mierocrystalline cellulose after processing in the fibrous material, the content of hemicellulose, measurement result is as shown in table 1;
The enzymic hydrolysis treatment step is identical with embodiment 1.
Enzymic hydrolysate is carried out centrifugal treating, get supernatant liquor after centrifugal and carry out reducing sugar content according to DNS method (dinitrosalicylic acid method) and measure, calculate Reducing sugar, measurement result is as shown in table 2.
Embodiment 3
In the pulverization process step of raw material except with water ratio be 5% Cortex Populi Tomentosae wood powder to be broken into granularity be 20-80 purpose powder, adding water, to make water ratio be that all the other are identical with embodiment 1 outside 75% the pre-treatment material;
The fungus degrading treatment step is 10:100 except the ratio of the volume of brown rot fungus inoculation suspension and the weight of pre-treatment material, i.e. the described brown rot fungus inoculation of inoculation 10L suspension in the described brown rot fungus inoculation suspension of inoculation 10ml or the every 100kg pre-treatment material in every 100g pre-treatment material; Culture temperature is 30 ℃, and incubation time is outside 20 days, and all the other are identical with embodiment 1;
The chemical degradation treatment step remains 180 ℃ except the temperature of reaction of reactor, DeR time 30min, tap water sprinkle reactor, until the temperature in the reactor drops to room temperature (35 ℃), residue after the cleaning carries out drying and processing under 40 ℃, drying time is 2 days, and the water ratio of the residue after the oven dry is outside 5%, all the other are identical with embodiment 1, measure the collaborative FeCl of fungi according to the standard method in U.S. renewable energy source laboratory
3Process the Mierocrystalline cellulose in the fibrous material, the content of hemicellulose, measurement result is as shown in table 1;
The enzymic hydrolysis treatment step is identical with embodiment 1.
Enzymic hydrolysate is carried out centrifugal treating, get supernatant liquor after centrifugal and carry out reducing sugar content and measure, measure according to DNS method (dinitrosalicylic acid method), measurement result is as shown in table 2.
Reference examples 1
With the Cortex Populi Tomentosae dry powder of embodiment 1-3 Raw pulverization process step preparation example 1 in contrast.
The Mierocrystalline cellulose in the fibrous material, the content of hemicellulose are measured in standard method according to U.S. renewable energy source laboratory, namely add mass percent concentration in the Cortex Populi Tomentosae powder and be 72% sulfuric acid, be under 30 ℃ in temperature, behind the hydrolysis 1h, adding deionized water, above-mentioned substance is diluted to concentration is 4% mixed liquor, then mixed liquor being put in the Autoclave, is under 121 ℃ the condition in temperature, heating 1h.Utilize poor heavy method to measure content of lignin.
Utilize high performance liquid phase ion chromatograph (Dai An company) to measure the Mierocrystalline cellulose in the unprocessed fibrous material, the content of hemicellulose.High-efficiency anion exchange ion chromatographic column (ICS3000 wears peace, the U.S.) is equipped with pulsed amperometric detector (CAD), AS50 automatic sampler, glycan analysis post (PA-20,4 * 250mm wear peace) and guard column (PA-20,3 * 30mm wear peace).The concrete analysis condition is: adopt the drip washing of 2.75mM NaOH isoconcentration to measure 6 kinds of contents of monosaccharides, flow velocity is 0.4mL/min, and the drip washing time is 40min.Adopt subsequently the drip washing of 1M NaAc isoconcentration to measure 2 kinds of glucuronic acid contents, flow velocity is 0.4mL/min, and the drip washing time is 10min.Adopt at last 137.4mM NaOH to clean chromatographic column 10min.The content of monose and uronic acid is obtained by the typical curve conversion of concentration known.Wherein the monose standard specimen is L-rhamnosyl, L-arabinose, D-Glucose, D-wood sugar, D-MANNOSE and D-semi-lactosi, and the standard specimen of uronic acid is D-Glucose aldehydic acid and D-galacturonic acid.Wherein, the content of sample fiber element is that content with glucose that this sample is surveyed multiply by hydrolysis coefficient 0.9 and obtains; The content of hemicellulose is the summation of wood sugar that this sample is surveyed, semi-lactosi and pectinose, and wherein wood sugar and galactose on hydrolysis coefficient are 0.88, and pectinose hydrolysis coefficient is 0.9.Measurement result is as shown in table 1.
Mierocrystalline cellulose, hemicellulose, content of lignin in table 1 biomass
| Plant fiber material | Mierocrystalline cellulose (g/g) | Hemicellulose (g/g) | Xylogen (g/g) |
| Embodiment 1 | 0.44 | 0.04 | 0.21 |
| Embodiment 2 | 0.33 | 0.007 | 0.22 |
| Embodiment 3 | 0.23 | 0 | 0.41 |
| The Cortex Populi Tomentosae raw material powder | 0.44 | 0.19 | 0.27 |
| Reference examples 2 | 0.43 | 0.06 | 0.21 |
| Reference examples 3 | 0.37 | 0.001 | 0.21 |
| Reference examples 4 | 0.37 | 0 | 0.26 |
| Reference examples 5 | 0.43 | 0.17 | 0.26 |
| Reference examples 6 | 0.41 | 0.15 | 0.21 |
| Reference examples 7 | 0.39 | 0.11 | 0.20 |
Show from the check result of table 1: rotten brown bacterium F.palustris and FeCl
3In 180 ℃ of collaborative pre-treatment Cortex Populi Tomentosae, wherein whole hemicelluloses is removed.In degradation of hemicellulose, when pretreatment temperature was higher than 140 ℃, Mierocrystalline cellulose also can be degraded, but trend is faint, so pretreatment temperature should be controlled at below 180 ℃.
Reference examples 2
1, chemical degradation is processed
Place by the cubic capacity of the stainless steel casting reactor (the mechanical company limited of the prosperous Sheng in Zhejiang) as 50mL take the Cortex Populi Tomentosae dry powder of embodiment 1-3 Raw pulverization process step preparation, then add the FeCl that concentration is 0.1mol/L
3Solution, the wherein weight of Cortex Populi Tomentosae dry powder and FeCl
3The ratio of the volume of solution is 10:100;
Reactor is placed the IKA magnetic force well heater (C-MAG HS7) that oil bath is installed, reactor is carried out the oil bath heating, so that the temperature in the reactor remains 140 ℃, fibrous material carries out chemical degradation reaction 30min, then take out reactor, be the tap water sprinkle reactor of (10 ℃) with temperature, until the temperature in the reactor drops to room temperature (25 ℃);
Filter the reaction degradation product in the reactor, then repeatedly clean undissolved residue with distilled water, until the purge flow fluid is neutral, namely the pH value of effluent liquid is 7; Then the residue after will cleaning carries out drying and processing under 35 ℃, drying time is 2 days, and the water ratio of the residue after the oven dry is about 7%, namely obtains FeCl
3The degradation of fibers raw material.
FeCl is measured in standard method according to U.S. renewable energy source laboratory
3Mierocrystalline cellulose in the degradation of fibers raw material, the content of hemicellulose, measurement result is as shown in table 1.
2, enzymic hydrolysis is processed
In 10mL sodium acetate buffer (50mM, pH4.8), add the tetracycline of 40 μ L and the cycloheximide solution of 20 μ L, mix, make the enzymic hydrolysis solvent, for subsequent use, wherein the concentration of tetracycline is 10%(m/v), the concentration of cycloheximide solution is 10% (m/v);
Add the 0.2g(dry weight to the enzymic hydrolysis solvent) FeCl
3Degradation of fibers raw material, cellulase and β-Portugal's glycosides carbohydrase stir to be placed on and carry out enzymic hydrolysis on the shaking table and process 96h and namely get enzymic hydrolysate, and its cellulase is that the 35FPU/g-substrate (is FeCl
3Preprocessing biomass), β-Portugal's glycosides carbohydrase is the 37.5IU/g-substrate, and the temperature that enzymic hydrolysis is processed is 50 ℃, and rotating speed is 150rpm.
Enzymic hydrolysate is carried out centrifugal treating, get supernatant liquor after centrifugal and carry out reducing sugar content and measure, measure reducing sugar content according to DNS method (dinitrosalicylic acid method), calculate Reducing sugar, the result is as shown in table 2.
Reference examples 3
Except the temperature of reactor in the chemical degradation treatment step remains 160 ℃, the DeR time is outside the 25min, and all the other are identical with reference examples 2, FeCl
3Mierocrystalline cellulose in the degradation of fibers raw material, the content of hemicellulose, measurement result is as shown in table 1; Through FeCl
3Reducing sugar content after the enzymic hydrolysis of chemical degradation fibrous material, Reducing sugar measurement result are as shown in table 2.
Reference examples 4
Except the temperature of reactor in the chemical degradation treatment step remains 180 ℃, the DeR time is outside the 30min, and all the other are identical with reference examples 3, FeCl
3Mierocrystalline cellulose in the degradation of fibers raw material, the content of hemicellulose, measurement result is as shown in table 1; Through FeCl
3Reducing sugar content after the enzymic hydrolysis of chemical degradation fibrous material, Reducing sugar measurement result are as shown in table 2.
Reference examples 5
Process except not carrying out chemical degradation, directly the pretreated fibrous material of fungi is carried out outside the enzymic hydrolysis, all the other are identical with embodiment 1, measure fungi according to the standard method in U.S. renewable energy source laboratory and process the Mierocrystalline cellulose in the rear fibrous material, the content of hemicellulose, measurement result is as shown in table 1; Reducing sugar content, Reducing sugar measurement result are as shown in table 2 after the enzymic hydrolysis of fungus degrading fibrous material.
Reference examples 6
Process except not carrying out chemical degradation, directly the pretreated fibrous material of fungi is carried out outside the enzymic hydrolysis, all the other are identical with embodiment 2, measure fungi according to the standard method in U.S. renewable energy source laboratory and process the Mierocrystalline cellulose in the rear fibrous material, the content of hemicellulose, measurement result is as shown in table 1; Reducing sugar content, Reducing sugar measurement result are as shown in table 2 after the enzymic hydrolysis of fungus degrading fibrous material.
Reference examples 7
Process except not carrying out chemical degradation, directly the pretreated fibrous material of fungi is carried out outside the enzymic hydrolysis, all the other are identical with embodiment 3, measure fungi according to the standard method in U.S. renewable energy source laboratory and process the Mierocrystalline cellulose in the rear fibrous material, the content of hemicellulose, measurement result is as shown in table 1; Reducing sugar content, Reducing sugar measurement result are as shown in table 2 after the enzymic hydrolysis of fungus degrading fibrous material.
Table 2 Reducing sugar measurement result
| ? | Reducing sugar (%) |
| Embodiment 1 | 26.63 |
| Embodiment 2 | 49.00 |
| Embodiment 3 | 95.36 |
| Reference examples 2 | 26.81 |
| Reference examples 3 | 36.93 |
| Reference examples 4 | 60.14 |
| Reference examples 5 | 20.81 |
| Reference examples 6 | 24.21 |
| Reference examples 7 | 25.61 |
The detected result of table 2 shows:
1, along with FeCl
3The rising of chemical degradation temperature (in 140-180 ℃ of scope), the content of reducing sugar is all in rising trend;
2, under uniform temp, adopt the Reducing sugar of the collaborative pre-treatment Cortex Populi Tomentosae of fungus degrading and chemical degradation high;
3, in the time of 180 ℃, adopt the collaborative pretreated Cortex Populi Tomentosae of fungus degrading and chemical degradation, after enzymic hydrolysis, the yield of reducing sugar reaches 95.36%, is independent FeCl under the equal conditions
3Pretreated 1.58 times.This illustrates the FeCl that rotten brown bacterium F.palustris is auxiliary
3Pre-treatment can increase cellulosic surface-area and porosity in the wood cell wall significantly, thereby increased Mierocrystalline cellulose to the accessibility of lytic enzyme and promoted enzymic hydrolysis efficient, causes the rising of Reducing sugar.
Test example 1X x ray diffraction analysis x
With X-ray diffractometer (Japanese Shimadzu company) among the embodiment 1-3 successively through brown rot fungus fungus degrading, FeCl
3FeCl among the Cortex Populi Tomentosae dry powder of the Cortex Populi Tomentosae powder of preparation, reference examples 1, the reference examples 2-4 after chemical degradation is processed
3Degraded Cortex Populi Tomentosae powder carries out X-ray diffraction, analyzes the degree of crystallinity of Cortex Populi Tomentosae powder, and the X-ray diffraction result as shown in Figure 1, 2.
X-ray diffraction result shown in Fig. 1,2 shows: the Cortex Populi Tomentosae powder is after the condition that is lower than 160 ℃ through temperature is processed, and the diffracted wave peak intensity of the X ray of sample can strengthen gradually; When temperature of reaction is higher than 160 ℃, the diffracted wave peak intensity of the X ray of sample will die down gradually.
Be lower than in temperature under 160 ℃ the treatment condition, the crystalline cellulose loss is faint, and a large amount of losses of noncrystalline domain hemicellulose, so the degree of crystallinity of sample can continue to rise, the diffracted wave peak intensity that shows as the X ray of sample strengthens; After temperature was higher than 160 ℃, because the minimizing of crystalline cellulose, sample degree of crystallinity began to reduce, so the diffracted wave peak intensity of X ray dies down.Therefore temperature be higher than process under 160 ℃ of conditions after, Mierocrystalline cellulose strengthens the accessibility of cellulase in the sample, thereby has improved enzymic hydrolysis efficient.
Test example 2 electron-microscope scanning analyses
To the collaborative FeCl of brown rot fungus among the Cortex Populi Tomentosae powder of brown rot fungus fungus degrading treatment step preparation among the embodiment 3, the embodiment 3
3FeCl in Cortex Populi Tomentosae powder after chemical hydrolysis is processed, the Cortex Populi Tomentosae dry powder of reference examples 1, the reference examples 4
3Degraded Cortex Populi Tomentosae powder adopts scanning electron microscope (HIT) to observe and takes the Cortex Populi Tomentosae dry powder, carry out electronic microscope photos, and electron-microscope scanning as shown in Figure 3.
Shown Cortex Populi Tomentosae dry powder (being untreated sample, reference examples 1) flat-satin by electron-microscope scanning result shown in Figure 3; Cortex Populi Tomentosae material sample (the embodiment 3 brown rot fungus degradation treatment Cortex Populi Tomentosae powder) surface of processing through brown rot fungus begins to have curling, and brown rot fungus degraded part hemicellulose and the cellulose components of Cortex Populi Tomentosae are described; Process directly employing FeCl without the fungi brown rot fungus
3Chemical degradation Cortex Populi Tomentosae material sample (reference examples 4) surface portion breaks, and demixing phenomenon is arranged, and FeCl is described
3Process the half a large amount of fibre fractionation of the Cortex Populi Tomentosae of having degraded in a large number; And the present invention adopts first the fungi pre-treatment, is then adopting FeCl
3Pretreated Cortex Populi Tomentosae sample is (among the embodiment 3 successively through brown rot fungus, FeCl
3Cortex Populi Tomentosae powder after the processing) skin breakage is serious, demixing phenomenon and aperture occur become many, the particle diameter of sample reduces, and has increased cellulosic surface-area and porosity in the wood cell wall, thereby increased the accessibility of Mierocrystalline cellulose to lytic enzyme, significantly promoted enzymolysis efficiency.
Claims (10)
1. a pretreatment process that improves the plant fiber material enzymolysis efficiency comprises that plant fiber material is carried out fungus degrading successively to be processed and FeCl
3Hydrolysis treatment.
2. pretreatment process as claimed in claim 1, it is characterized in that described fungus degrading process be with plant fiber material with after the fungi inoculation liquid is mixed, be under 28 ± 3 ℃ in temperature, cultivated 20-40 days.
3. pretreatment process as claimed in claim 2 is characterized in that described fungi selection brown rot fungus (Fomitopsis palustris C7615).
4. pretreatment process as claimed in claim 1 or 2 is characterized in that described plant fiber material is prepared from as follows: at first plant fiber material is pulverized; Then the water ratio of regulating plant fibrous material is to 70-80%; Then plant fiber material is carried out sterilising treatment, and get final product.
5. pretreatment process as claimed in claim 4, the granularity of the plant fiber material after it is characterized in that pulverizing is the 20-80 order.
6. pretreatment process as claimed in claim 2, it is characterized in that described fungi inoculation liquid is prepared from accordance with the following steps: at first fungi is inoculated in and amplifies in substratum, under dark condition, in 28 ± 1 ℃, rotating speed is to cultivate 3-7 days under the condition of 100-150rpm, then add sterile distilled water, carry out homogenized, and get final product.
7. pretreatment process as claimed in claim 6, it is characterized in that the composition of the described amplification culture base of every 1000ml consists of: 20g glucose, 5g yeast soak powder, 1g KH
2PO
4, 0.5g MgSO
4, the 0.01g vitamins B
1, it is 1000ml that deionized water is regulated cumulative volume.
8. pretreatment process as claimed in claim 1 is characterized in that described FeCl
3Hydrolysis treatment comprise following in sequence step:
A) with plant fiber material and FeCl after the fungus degrading processing
3Solution mixes, and makes FeCl
3The hydrolysis mixed solution;
B) heating FeCl
3The hydrolysis mixed solution is the reaction 20-30min that is hydrolyzed under 140-180 ℃ keeping temperature;
C) to the processing of lowering the temperature of the mixture behind the hydrolysis reaction, until the temperature of hydrolysis reaction mixture reaches 15-35 ℃.
9. pretreatment process as claimed in claim 8 is characterized in that steps A) described in weight and the FeCl of the plant fiber material of fungus degrading after processing
3The ratio of the volume of solution is 7.5-15:10.
10. pretreatment process as claimed in claim 8 is characterized in that steps A) described in FeCl
3The concentration of solution is 0.05-0.2mol/L.
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110283334A (en) * | 2019-07-02 | 2019-09-27 | 哈尔滨理工大学 | A kind of preprocess method improving ash biodegrade effect |
| CN110747236A (en) * | 2018-07-23 | 2020-02-04 | 天津市职业大学 | Pretreatment method of lignocellulose and application thereof in improving lignocellulose enzymolysis efficiency |
| CN113183266A (en) * | 2021-04-27 | 2021-07-30 | 中南林业科技大学 | Method for pretreating poplar fibers by hydrothermal-peroxyacetate metal salt cooperation |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1178077A (en) * | 1997-10-24 | 1998-04-08 | 天津市农业生物工程研究中心 | Biogenic additives for fermentation of stalks and preparation thereof |
| CN101486988A (en) * | 2009-02-12 | 2009-07-22 | 中国农业大学 | Method for degrading ligno-cellulose and special cellulose decomposing bacteria therefor |
| CN102517947A (en) * | 2011-12-21 | 2012-06-27 | 张健 | Production method for paper pulp by using brown rot fungus in biodegradation of lignin |
-
2012
- 2012-12-31 CN CN2012105942359A patent/CN103014094A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1178077A (en) * | 1997-10-24 | 1998-04-08 | 天津市农业生物工程研究中心 | Biogenic additives for fermentation of stalks and preparation thereof |
| CN101486988A (en) * | 2009-02-12 | 2009-07-22 | 中国农业大学 | Method for degrading ligno-cellulose and special cellulose decomposing bacteria therefor |
| CN102517947A (en) * | 2011-12-21 | 2012-06-27 | 张健 | Production method for paper pulp by using brown rot fungus in biodegradation of lignin |
Non-Patent Citations (1)
| Title |
|---|
| 王蔚等: "褐腐真菌纤维素降解初期产生羟基自由基的普遍性研究", 《菌物系统》 * |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110747236A (en) * | 2018-07-23 | 2020-02-04 | 天津市职业大学 | Pretreatment method of lignocellulose and application thereof in improving lignocellulose enzymolysis efficiency |
| CN110283334A (en) * | 2019-07-02 | 2019-09-27 | 哈尔滨理工大学 | A kind of preprocess method improving ash biodegrade effect |
| CN110283334B (en) * | 2019-07-02 | 2021-07-23 | 哈尔滨理工大学 | Pretreatment method for improving biodegradation effect of locust wood |
| CN113183266A (en) * | 2021-04-27 | 2021-07-30 | 中南林业科技大学 | Method for pretreating poplar fibers by hydrothermal-peroxyacetate metal salt cooperation |
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