CN111733197A - Method for enzymatic hydrolysis and saccharification of plant fiber raw material - Google Patents
Method for enzymatic hydrolysis and saccharification of plant fiber raw material Download PDFInfo
- Publication number
- CN111733197A CN111733197A CN202010616097.4A CN202010616097A CN111733197A CN 111733197 A CN111733197 A CN 111733197A CN 202010616097 A CN202010616097 A CN 202010616097A CN 111733197 A CN111733197 A CN 111733197A
- Authority
- CN
- China
- Prior art keywords
- plant fiber
- saccharification
- fiber raw
- enzyme
- enzymatic hydrolysis
- 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.)
- Pending
Links
- 239000002994 raw material Substances 0.000 title claims abstract description 30
- 239000000835 fiber Substances 0.000 title claims abstract description 29
- 238000000034 method Methods 0.000 title claims abstract description 29
- 230000007071 enzymatic hydrolysis Effects 0.000 title claims abstract description 25
- 238000006047 enzymatic hydrolysis reaction Methods 0.000 title claims abstract description 25
- 108090000790 Enzymes Proteins 0.000 claims abstract description 33
- 102000004190 Enzymes Human genes 0.000 claims abstract description 33
- 229940088598 enzyme Drugs 0.000 claims abstract description 33
- 230000007062 hydrolysis Effects 0.000 claims abstract description 11
- 238000006460 hydrolysis reaction Methods 0.000 claims abstract description 11
- 108010059892 Cellulase Proteins 0.000 claims abstract description 9
- 229940106157 cellulase Drugs 0.000 claims abstract description 9
- 238000000926 separation method Methods 0.000 claims abstract description 9
- 239000007853 buffer solution Substances 0.000 claims abstract description 6
- 241000196324 Embryophyta Species 0.000 claims description 27
- 239000007788 liquid Substances 0.000 claims description 15
- 239000001913 cellulose Substances 0.000 claims description 14
- 229920002678 cellulose Polymers 0.000 claims description 14
- 230000008569 process Effects 0.000 claims description 11
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 9
- 239000000413 hydrolysate Substances 0.000 claims description 8
- 235000017166 Bambusa arundinacea Nutrition 0.000 claims description 6
- 235000017491 Bambusa tulda Nutrition 0.000 claims description 6
- 235000015334 Phyllostachys viridis Nutrition 0.000 claims description 6
- 239000011425 bamboo Substances 0.000 claims description 6
- 230000009849 deactivation Effects 0.000 claims description 6
- 101710121765 Endo-1,4-beta-xylanase Proteins 0.000 claims description 5
- 241001520808 Panicum virgatum Species 0.000 claims description 5
- 108010059820 Polygalacturonase Proteins 0.000 claims description 5
- 230000002255 enzymatic effect Effects 0.000 claims description 5
- 108010093305 exopolygalacturonase Proteins 0.000 claims description 5
- 108010084650 alpha-N-arabinofuranosidase Proteins 0.000 claims description 3
- 241000209140 Triticum Species 0.000 claims description 2
- 235000021307 Triticum Nutrition 0.000 claims description 2
- 240000008042 Zea mays Species 0.000 claims description 2
- 235000005824 Zea mays ssp. parviglumis Nutrition 0.000 claims description 2
- 235000002017 Zea mays subsp mays Nutrition 0.000 claims description 2
- 235000005822 corn Nutrition 0.000 claims description 2
- 238000001125 extrusion Methods 0.000 claims description 2
- 238000004537 pulping Methods 0.000 claims description 2
- 238000010298 pulverizing process Methods 0.000 claims 3
- 240000003433 Miscanthus floridulus Species 0.000 claims 1
- 244000082204 Phyllostachys viridis Species 0.000 claims 1
- 239000010907 stover Substances 0.000 claims 1
- 239000010902 straw Substances 0.000 claims 1
- 229920005610 lignin Polymers 0.000 abstract description 13
- 239000000758 substrate Substances 0.000 abstract description 13
- 239000002028 Biomass Substances 0.000 abstract description 10
- 239000003153 chemical reaction reagent Substances 0.000 abstract description 5
- 239000007787 solid Substances 0.000 abstract description 3
- 238000006243 chemical reaction Methods 0.000 abstract description 2
- 238000006116 polymerization reaction Methods 0.000 abstract description 2
- 239000001509 sodium citrate Substances 0.000 abstract description 2
- NLJMYIDDQXHKNR-UHFFFAOYSA-K sodium citrate Chemical compound O.O.[Na+].[Na+].[Na+].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O NLJMYIDDQXHKNR-UHFFFAOYSA-K 0.000 abstract description 2
- VMHLLURERBWHNL-UHFFFAOYSA-M Sodium acetate Chemical compound [Na+].CC([O-])=O VMHLLURERBWHNL-UHFFFAOYSA-M 0.000 abstract 1
- 239000001632 sodium acetate Substances 0.000 abstract 1
- 235000017281 sodium acetate Nutrition 0.000 abstract 1
- 239000000463 material Substances 0.000 description 15
- 210000002421 cell wall Anatomy 0.000 description 12
- 241000878006 Miscanthus sinensis Species 0.000 description 11
- 241000878007 Miscanthus Species 0.000 description 6
- 241001330002 Bambuseae Species 0.000 description 5
- 238000005516 engineering process Methods 0.000 description 5
- 239000000203 mixture Substances 0.000 description 5
- QGZKDVFQNNGYKY-UHFFFAOYSA-N Ammonia Chemical compound N QGZKDVFQNNGYKY-UHFFFAOYSA-N 0.000 description 4
- 238000006731 degradation reaction Methods 0.000 description 4
- QTBSBXVTEAMEQO-UHFFFAOYSA-N Acetic acid Chemical compound CC(O)=O QTBSBXVTEAMEQO-UHFFFAOYSA-N 0.000 description 3
- 230000004888 barrier function Effects 0.000 description 3
- 230000015556 catabolic process Effects 0.000 description 3
- 238000001035 drying Methods 0.000 description 3
- 230000007613 environmental effect Effects 0.000 description 3
- 238000000227 grinding Methods 0.000 description 3
- 238000003306 harvesting Methods 0.000 description 3
- 150000002772 monosaccharides Chemical class 0.000 description 3
- 238000007670 refining Methods 0.000 description 3
- 239000000243 solution Substances 0.000 description 3
- 238000005406 washing Methods 0.000 description 3
- RGHNJXZEOKUKBD-SQOUGZDYSA-N D-gluconic acid Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@@H](O)C(O)=O RGHNJXZEOKUKBD-SQOUGZDYSA-N 0.000 description 2
- QXKAIJAYHKCRRA-UHFFFAOYSA-N D-lyxonic acid Natural products OCC(O)C(O)C(O)C(O)=O QXKAIJAYHKCRRA-UHFFFAOYSA-N 0.000 description 2
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 2
- YLQBMQCUIZJEEH-UHFFFAOYSA-N Furan Chemical compound C=1C=COC=1 YLQBMQCUIZJEEH-UHFFFAOYSA-N 0.000 description 2
- 241000589232 Gluconobacter oxydans Species 0.000 description 2
- 229920002488 Hemicellulose Polymers 0.000 description 2
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 description 2
- LRHPLDYGYMQRHN-UHFFFAOYSA-N N-Butanol Chemical compound CCCCO LRHPLDYGYMQRHN-UHFFFAOYSA-N 0.000 description 2
- 229910021529 ammonia Inorganic materials 0.000 description 2
- 230000003197 catalytic effect Effects 0.000 description 2
- 238000004880 explosion Methods 0.000 description 2
- HYBBIBNJHNGZAN-UHFFFAOYSA-N furfural Chemical compound O=CC1=CC=CO1 HYBBIBNJHNGZAN-UHFFFAOYSA-N 0.000 description 2
- 238000012239 gene modification Methods 0.000 description 2
- 230000005017 genetic modification Effects 0.000 description 2
- 235000013617 genetically modified food Nutrition 0.000 description 2
- 230000006872 improvement Effects 0.000 description 2
- JVTAAEKCZFNVCJ-UHFFFAOYSA-N lactic acid Chemical compound CC(O)C(O)=O JVTAAEKCZFNVCJ-UHFFFAOYSA-N 0.000 description 2
- 238000004519 manufacturing process Methods 0.000 description 2
- 244000005700 microbiome Species 0.000 description 2
- 238000003801 milling Methods 0.000 description 2
- 239000000178 monomer Substances 0.000 description 2
- 238000002203 pretreatment Methods 0.000 description 2
- 239000007974 sodium acetate buffer Substances 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- KDYFGRWQOYBRFD-UHFFFAOYSA-N succinic acid Chemical compound OC(=O)CCC(O)=O KDYFGRWQOYBRFD-UHFFFAOYSA-N 0.000 description 2
- 239000002351 wastewater Substances 0.000 description 2
- 241000228245 Aspergillus niger Species 0.000 description 1
- RGHNJXZEOKUKBD-UHFFFAOYSA-N D-gluconic acid Natural products OCC(O)C(O)C(O)C(O)C(O)=O RGHNJXZEOKUKBD-UHFFFAOYSA-N 0.000 description 1
- QXKAIJAYHKCRRA-FLRLBIABSA-N D-xylonic acid Chemical compound OC[C@@H](O)[C@H](O)[C@@H](O)C(O)=O QXKAIJAYHKCRRA-FLRLBIABSA-N 0.000 description 1
- RGHNJXZEOKUKBD-KLVWXMOXSA-N L-gluconic acid Chemical compound OC[C@H](O)[C@H](O)[C@@H](O)[C@H](O)C(O)=O RGHNJXZEOKUKBD-KLVWXMOXSA-N 0.000 description 1
- QXKAIJAYHKCRRA-NUNKFHFFSA-N L-xylonic acid Chemical compound OC[C@H](O)[C@@H](O)[C@H](O)C(O)=O QXKAIJAYHKCRRA-NUNKFHFFSA-N 0.000 description 1
- 238000010306 acid treatment Methods 0.000 description 1
- 238000007605 air drying Methods 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 238000000498 ball milling Methods 0.000 description 1
- 108010047754 beta-Glucosidase Proteins 0.000 description 1
- 102000006995 beta-Glucosidase Human genes 0.000 description 1
- 239000006227 byproduct Substances 0.000 description 1
- 238000004364 calculation method Methods 0.000 description 1
- 238000006555 catalytic reaction Methods 0.000 description 1
- 210000004027 cell Anatomy 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 238000010276 construction Methods 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 230000018044 dehydration Effects 0.000 description 1
- 238000006297 dehydration reaction Methods 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000005265 energy consumption Methods 0.000 description 1
- 238000003912 environmental pollution Methods 0.000 description 1
- 238000000855 fermentation Methods 0.000 description 1
- 230000004151 fermentation Effects 0.000 description 1
- 238000004108 freeze drying Methods 0.000 description 1
- 239000000174 gluconic acid Substances 0.000 description 1
- 235000012208 gluconic acid Nutrition 0.000 description 1
- 229940059442 hemicellulase Drugs 0.000 description 1
- 108010002430 hemicellulase Proteins 0.000 description 1
- 239000003112 inhibitor Substances 0.000 description 1
- 239000004310 lactic acid Substances 0.000 description 1
- 235000014655 lactic acid Nutrition 0.000 description 1
- 238000002386 leaching Methods 0.000 description 1
- 230000002503 metabolic effect Effects 0.000 description 1
- 230000000813 microbial effect Effects 0.000 description 1
- 210000001724 microfibril Anatomy 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 229920001542 oligosaccharide Polymers 0.000 description 1
- 150000002482 oligosaccharides Chemical class 0.000 description 1
- 239000003960 organic solvent Substances 0.000 description 1
- 239000002245 particle Substances 0.000 description 1
- 150000002989 phenols Chemical class 0.000 description 1
- 229920000642 polymer Polymers 0.000 description 1
- 238000002390 rotary evaporation Methods 0.000 description 1
- 239000001384 succinic acid Substances 0.000 description 1
- 210000001519 tissue Anatomy 0.000 description 1
- 231100000331 toxic Toxicity 0.000 description 1
- 230000002588 toxic effect Effects 0.000 description 1
- 239000011882 ultra-fine particle Substances 0.000 description 1
Images
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
- C12P19/00—Preparation of compounds containing saccharide radicals
- C12P19/14—Preparation of compounds containing saccharide radicals produced by the action of a carbohydrase (EC 3.2.x), e.g. by alpha-amylase, e.g. by cellulase, hemicellulase
-
- 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
- C12P19/00—Preparation of compounds containing saccharide radicals
- C12P19/02—Monosaccharides
-
- 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
- C12P2201/00—Pretreatment of cellulosic or lignocellulosic material for subsequent enzymatic treatment or hydrolysis
-
- 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
- C12P2203/00—Fermentation products obtained from optionally pretreated or hydrolyzed cellulosic or lignocellulosic material as the carbon source
Landscapes
- Organic Chemistry (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Zoology (AREA)
- Life Sciences & Earth Sciences (AREA)
- Wood Science & Technology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Microbiology (AREA)
- General Chemical & Material Sciences (AREA)
- Biotechnology (AREA)
- Health & Medical Sciences (AREA)
- Biochemistry (AREA)
- Bioinformatics & Cheminformatics (AREA)
- General Engineering & Computer Science (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
The invention discloses a method for enzymatic hydrolysis and saccharification of plant fiber raw materials, and belongs to the technical field of biomass separation and conversion. The method comprises the following steps: mechanically treating fresh plant fiber raw materials at the early growth and development stage to obtain disintegrated plant fiber raw materials; adding cellulase and the like into a buffer solution of sodium citrate or sodium acetate by taking the crushed plant fiber raw material as a substrate to carry out biological enzyme hydrolysis; due to the fact that the lignification degree of the fresh plant fibers in the early growth stage is low, the lignin polymerization degree is also low, the accessibility of the enzymes to the fibers is high, and reasonable mechanical treatment is added, the method provided by the invention can promote the enzymatic hydrolysis saccharification efficiency of the plant fiber raw materials to be as high as 95.5% under the condition that a large number of chemical reagents are not used, meanwhile, the obtained solid residues rich in the primary lignin components can be further utilized in a high-value mode, and the method is green, efficient and high in application and popularization values.
Description
Technical Field
The invention belongs to the technical field of biomass separation and conversion, and particularly relates to a method for enzymatic hydrolysis and saccharification of a plant fiber raw material.
Background
With the rapid development of economy in China, the contradiction between energy supply and demand and the environmental problem will become increasingly serious. Therefore, the full, effective and scientific development and utilization of biomass energy is one of the ways for relieving the energy contradiction in China in the future. The development of biomass energy not only relieves the energy shortage and environmental pressure of China, but also can promote rural economy, promote the income increase of farmers and promote new rural construction, and meanwhile, the whole field of biomass energy is listed in an encouragement catalog by national development and improvement committee in this year.
Cellulose and hemicellulose in the lignocellulose raw material can be degraded into monosaccharide under the catalysis of biological enzyme, and the monosaccharide can be fermented by microorganisms to produce biological energy sources such as ethanol and butanol and high-valued platform compounds such as lactic acid and succinic acid. And the plant forms a complex cell wall structure in the growth and development process, the lignification degree of the fiber cell wall is increased and the fiber crystallinity is gradually improved along with the growth and development of the plant, the accessibility of the biological enzyme is reduced, so that the cell wall is difficult to be degraded by the enzyme, namely a 'biomass anti-degradation barrier', and the biorefinery difficulty is increased. Therefore, the biorefinery of the lignocellulose raw material needs to break the biomass degradation-resistant barrier through a pretreatment link, and the mainly adopted pretreatment technologies comprise dilute acid treatment, steam explosion, Ammonia Fiber Explosion (AFEX), low-temperature ammonia leaching treatment, high-temperature liquid water treatment, an organic solvent method, an alkaline hydrogen peroxide method and the like, wherein the AFEX is one of the most effective methods for breaking the biomass degradation-resistant barrier and is also the most economical and feasible physical-chemical pretreatment technology, but the AFEX pretreatment has a limited effect on the plant fiber raw material with high lignin content. In addition, most pretreatment methods hydrolyze cell wall polymers to a certain extent, the generated oligosaccharides or monosaccharides are dissolved in pretreatment liquid or washing liquid and run off along with the treatment process, and in addition, byproducts such as acetic acid, furan, phenols and furfural are generated from hemicellulose and lignin in the process, so that the metabolic activity of microorganisms is inhibited. Therefore, the search and development of pretreatment methods with low cost, high efficiency, less sugar loss and less inhibitors are still the direction of further industrialization of the bio-refining technology of plant fiber resources.
Because the pretreatment means reported at present usually has the defects of harsh conditions, use of toxic and harmful reagents and the like, the energy consumption is high, the environmental pollution is serious, and the subsequent microbial fermentation is inhibited, experts in the field of biology find that the enzymolysis sugar production efficiency of the cell wall is determined structurally by researching the relation between key structural factors of the cell wall and the efficient degradation of biomass, so that cell wall genetic modification engineering technologies are provided, such as strategies of reducing the lignin content of the cell wall, changing the composition and the structure of lignin monomers, modifying the lignin monomers, increasing the low-crystallinity cellulose content of the cell wall, reducing the high-crystallinity cellulose content of the cell wall and the like. However, by utilizing transgenosis and mutants, the cell wall structure and composition of the mutants are changed systematically, so that the normal growth and development of plants are influenced, the yield of seeds and biomass is greatly changed, the period is long, the cost is high, and no breakthrough progress is obtained.
Research and analysis suggest that along with the growth and development of plants, the lignification degree of fibers is continuously increased, the polymerization degree of lignin is higher and higher, and the accessibility of enzymes to the fibers is reduced; the cohesion of the microfibril layer of the fiber structure is stronger and higher, the crystallinity is higher and higher, and the enzymatic hydrolysis and saccharification are more and more difficult; in addition, after the fresh plants are air-dried, the cohesion of the intercellular layer of the fiber cells is enhanced and the tissues are keratinized due to dehydration, so that the accessibility of the cellulase is weakened; the method has the advantages that the biological refining must adopt a pretreatment means or a genetic modification engineering technology to reduce the lignin content, change the lignin structure and reduce the cellulose crystallinity, thereby increasing the accessibility of the biological enzyme and improving the enzymatic hydrolysis saccharification efficiency, not only the process is complex and the biological refining cost is increased, but also a large amount of waste water is generated and a new environmental problem is formed.
Therefore, there is a need for a green and efficient enzymatic hydrolysis and saccharification method for fresh plant fiber raw material, which can promote the enzymatic hydrolysis and saccharification efficiency of lignocellulose raw material without using a large amount of chemical reagents, and can further utilize the separated native lignin in high value.
Disclosure of Invention
Aiming at the problems in the prior art, the technical problem to be solved by the invention is to provide a method for enzymatic hydrolysis and saccharification of plant fiber raw materials, which solves the problems that the existing cellulose enzymatic hydrolysis method is complex in process, uses a large amount of chemical reagents, generates a large amount of waste water and affects the environment and the like.
In order to solve the technical problems, the technical scheme adopted by the invention is as follows:
a method for enzymatic hydrolysis saccharification of plant fiber raw materials, comprising: fresh tender plants with rich cellulose content are used as raw materials, after deactivation of enzymes, mechanical crushing and moisture balancing, buffer solution and enzyme are added for hydrolysis, and enzyme hydrolysate is obtained through solid-liquid separation.
Further, the method for enzymatic hydrolysis and saccharification of the plant fiber raw material comprises the following specific steps:
1) obtaining fresh tender plants with rich cellulose content, immediately deactivating enzyme, completely removing leaves and sheaths, and reserving stem parts;
2) adding water into the stalks and then mechanically crushing the stalks to obtain crushed stalks;
3) balancing water content of the crushed stalks, and adding a buffer solution and enzyme for hydrolysis;
4) and (4) performing solid-liquid separation on the hydrolysate to obtain enzyme hydrolysate.
Further, the fresh tender plant rich in cellulose is one or more of miscanthus, switchgrass, bamboo, corn stalks and wheat stalks.
Furthermore, the fresh tender plant rich in cellulose has a growth time of 10 days to 6 months.
Further, the specific conditions of the fixation are as follows: 105 ℃ for 20 min.
Further, the mechanical crushing is one or a combination of more of wall breaking machine crushing, pulping by Bulboli, superfine particle crushing, disc milling, PFI milling, ball milling and double-screw extrusion.
Further, the enzyme is one or more of cellulase, xylanase, pectinase and arabinosidase.
Drawings
FIG. 1 is a diagram of fresh miscanthus after the leaves and sheaths are removed; in the figure, the left figure is the miscanthus comprising leaves and sheaths, the middle figure is the miscanthus with leaves and sheaths removed, and the right figure is the miscanthus with leaves and sheaths removed simultaneously;
FIG. 2 is a graph showing the comparison of the enzyme hydrolysis yield of fresh miscanthus sinensis at 96h before and after the fresh miscanthus sinensis is air-dried for 2, 4 and 6 months after growth and development; in the figure, FG: air-drying Chinese silvergrass, XX: fresh miscanthus;
FIG. 3 is a graph showing the results of producing gluconic acid and xylonic acid by using a whole-cell catalytic enzymatic hydrolysate of gluconobacter oxydans.
Detailed Description
The invention is further described with reference to specific examples.
The cellulase is Novoxil second-generation commercial enzyme (
CTec2, Novozymes), is an enzyme mixture comprising cellulase, β -glucosidase and hemicellulase xylanase (CAS9025-57-4), pectinase (CAS9032-75-1), derived from aspergillus niger, available from michelin (shanghai) inc.
Example 1:
fresh miscanthus sinensis with growth development period of 2 months, 4 months and 6 months is used as raw material, and water is removed immediately after harvesting (105 ℃, 20 min). Removing leaves, sheaths and other parts of the materials completely after completing the enzyme deactivation, only keeping the stem part (as shown in figure 1), and crushing for 15 times by a wall breaking machine under the condition that the substrate concentration is 10 percent (the mass ratio of stems to water is 1: 9), and obtaining crushed feed liquid 50s each time. Then carrying out disc grinding for 10 times by virtue of an ultrafine particle grinder under the conditions that the disc grinding gap is 200 mu m and the substrate concentration is 2 percent to obtain a crushed miscanthus sinensis material, placing the crushed miscanthus sinensis material in a refrigerator with the temperature of 4 ℃ for balancing moisture for 24h, measuring the moisture content of the material, drying a small amount of crushed material at room temperature, measuring the component content of the material, and obtaining the air-dried miscanthus sinensis raw material. Then, the mixture was added to 50mM sodium citrate buffer solution, pH4.8, at a solid-to-liquid ratio of 2% (w/v), and then, 50FPU/g substrate cellulase, 50FPU/g substrate xylanase and 50FPU/g substrate pectinase were added for enzymatic hydrolysis, and the mixture was subjected to enzymatic hydrolysis at 50 ℃ for 96 hours on a shaker at 150 rpm.
The results are shown in figure 2, and the enzyme hydrolysis yield of the fresh miscanthus sinensis for 96h in 2 months, 4 months and 6 months of growth and development is 95.5%, 62.11% and 25.66% respectively. And the enzyme hydrolysis yield of 96h of the air-dried miscanthus sinensis for 2 months, 4 months and 6 months of growth and development is 88.36%, 54.22% and 15.20% respectively. Compared with the mature air-dried Chinese silvergrass, the lignification degree of the fresh Chinese silvergrass raw material in the early growth and development stage is low, the cell wall structure can be damaged after the simple mechanical treatment is adopted, and the high-efficiency enzymatic hydrolysis saccharification of cellulose can be realized under the condition of not using a large amount of chemical reagents.
The calculation formula of the enzyme hydrolysis yield is shown as follows:
and performing solid-liquid separation on the enzymatic hydrolysate to obtain sugar liquor and solid residues rich in lignin, dialyzing and purifying the solid residues, and performing freeze drying to obtain primary lignin which can be further utilized at a high value. The sugar solution is concentrated by rotary evaporation until the total sugar concentration is about 100g/L, and 67.14g/L gluconic acid and 28.09g/L xylonic acid can be obtained within 24h by utilizing gluconobacter oxydans whole-cell catalytic production platform chemicals (as shown in figure 3).
Example 2:
fresh bamboo grown for 10 days and 50 days was used as a raw material, and immediately deactivated after harvesting (105 ℃, 20 min). Removing branches and leaves of the material after completing enzyme deactivation, only reserving a stem part, extruding once by using a double screw, and then grinding for 5 times by using a disc grinder, wherein the disc grinder interval is 1mm, obtaining a crushed bamboo raw material through solid-liquid separation and water washing processes, placing the crushed bamboo raw material in a refrigerator at 4 ℃ for balancing moisture for 24 hours, measuring the moisture content of the material, taking a small amount of crushed material, drying the crushed material at room temperature, and measuring the component content of the material. Then, 50FPU/g substrate cellulase, 20FPU/g substrate xylanase, 20FPU/g substrate arabinosidase and 20FPU/g substrate pectinase were added to 50mM sodium acetate buffer solution (pH 4.8) at a solid-to-liquid ratio of 2% (w/v) to carry out enzymatic hydrolysis, and the enzymatic hydrolysis was carried out at 50 ℃ for 96 hours on a shaker at 150 rpm. The yield of the bamboo enzyme hydrolysis of 10 days and 50 days of growth and development is 52.05% and 16.77% respectively (the formula of the enzyme hydrolysis yield is the same as that of example 1).
Example 3:
fresh switchgrass growing for 3 months is used as a raw material, and the enzyme is deactivated immediately after harvesting (105 ℃, 20 min). Removing branches and leaves of the material after completing the enzyme deactivation, only keeping the stem part, crushing for 10 times by a wall breaking machine under the condition that the substrate concentration is 10%, obtaining crushed feed liquid for 50s each time, then treating for 1h by a Waley beater under the substrate concentration of 2%, obtaining crushed switchgrass raw material through the processes of solid-liquid separation and water washing, placing in a refrigerator at 4 ℃ for balancing moisture for 24h, measuring the moisture content of the material, taking a small amount of crushed material, drying under the room temperature condition, and measuring the content of the material components. Then, 2% (w/v) of the solid-to-liquid ratio was added to 50mM sodium acetate buffer solution, pH4.8, and 50FPU/g of substrate cellulase was added to carry out the enzymatic hydrolysis at 50 ℃ for 96 hours on a shaker at 150 rpm. The enzymatic hydrolysis yield of switchgrass 3 months after growth was 51.79% (the formula for the enzymatic hydrolysis yield is the same as in example 1).
The above description is only for the purpose of illustrating the preferred embodiments of the present invention and is not to be construed as limiting the invention, and any modifications, equivalents and improvements made within the spirit and principle of the present invention are intended to be included within the scope of the present invention.
Claims (7)
1. A method for enzymatic hydrolysis and saccharification of plant fiber raw materials is characterized in that fresh tender plants with rich cellulose content are used as raw materials, after deactivation of enzymes, mechanical crushing and moisture balancing, buffer solution and enzyme are added for hydrolysis, and enzymatic hydrolysate is obtained through solid-liquid separation.
2. The method for enzymatic hydrolysis and saccharification of a plant fiber raw material according to claim 1, characterized by the specific steps of:
1) obtaining fresh tender plants with rich cellulose content, immediately deactivating enzyme, completely removing leaves and sheaths, and reserving stem parts;
2) adding water into the stalks and then mechanically crushing the stalks to obtain crushed stalks;
3) balancing water content of the crushed stalks, and adding a buffer solution and enzyme for hydrolysis;
4) and (4) performing solid-liquid separation on the hydrolysate to obtain enzyme hydrolysate.
3. The process of enzymatic hydrolysis saccharification of plant fiber feedstock as claimed in claim 1 or 2, wherein said fresh tender plant rich in cellulose is one or more of miscanthus, switchgrass, bamboo, corn stover and wheat straw.
4. The process for enzymatic saccharification of plant fiber feedstock as claimed in claim 1 or 2, wherein the fresh young plants rich in cellulose are grown for a period of 10 days to 6 months.
5. The process for enzymatic saccharification of plant fiber raw material as claimed in claim 1 or 2, wherein the specific conditions for deactivation of enzyme are: 105 ℃ for 20 min.
6. The process of enzymatic hydrolysis saccharification of plant fiber raw material as claimed in claim 1 or 2, characterized in that, the mechanical pulverization is one or more of breaking wall machine pulverization, pulping by Bulboro, ultramicron pulverization, disc mill, PFI mill, ball mill, twin screw extrusion.
7. The process according to claim 1 or 2, wherein the enzyme is a cellulase, xylanase, pectinase, arabinosidase or a combination thereof.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010616097.4A CN111733197A (en) | 2020-06-30 | 2020-06-30 | Method for enzymatic hydrolysis and saccharification of plant fiber raw material |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010616097.4A CN111733197A (en) | 2020-06-30 | 2020-06-30 | Method for enzymatic hydrolysis and saccharification of plant fiber raw material |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN111733197A true CN111733197A (en) | 2020-10-02 |
Family
ID=72653833
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202010616097.4A Pending CN111733197A (en) | 2020-06-30 | 2020-06-30 | Method for enzymatic hydrolysis and saccharification of plant fiber raw material |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN111733197A (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114293393A (en) * | 2021-12-24 | 2022-04-08 | 南京工业大学 | Process for improving biomass enzymolysis efficiency by combining alkali and double-screw extrusion pretreatment with biological additive |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105803017A (en) * | 2016-04-14 | 2016-07-27 | 南京林业大学 | Method for improving wood fiber material enzyme hydrolysis saccharification efficiency |
| CN107164424A (en) * | 2017-07-12 | 2017-09-15 | 天津科技大学 | It is a kind of to aoxidize the lignocellulose pretreatment method that delignification improves enzyme hydrolysis rate |
-
2020
- 2020-06-30 CN CN202010616097.4A patent/CN111733197A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105803017A (en) * | 2016-04-14 | 2016-07-27 | 南京林业大学 | Method for improving wood fiber material enzyme hydrolysis saccharification efficiency |
| CN107164424A (en) * | 2017-07-12 | 2017-09-15 | 天津科技大学 | It is a kind of to aoxidize the lignocellulose pretreatment method that delignification improves enzyme hydrolysis rate |
Non-Patent Citations (2)
| Title |
|---|
| HONGLIANG GUO 等: "Enzymatic saccharification of lignocellulosic biorefinery: Research focuses", 《BIORESOUR TECHNOL》 * |
| 侣胜利: "能源植物芒草细胞壁结构组成与糖化发酵关系的研究", 《中国博士学位论文全文数据库》 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114293393A (en) * | 2021-12-24 | 2022-04-08 | 南京工业大学 | Process for improving biomass enzymolysis efficiency by combining alkali and double-screw extrusion pretreatment with biological additive |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| Li et al. | Simultaneous saccharification and fermentation of lignocellulosic residues pretreated with phosphoric acid–acetone for bioethanol production | |
| US7754456B2 (en) | Process for producing ethanol | |
| Saratale et al. | Screening and optimization of pretreatments in the preparation of sugarcane bagasse feedstock for biohydrogen production and process optimization | |
| US20100159521A1 (en) | Ozone treatment of biomass to enhance enzymatic saccharification | |
| CN101514349B (en) | Method for preparing fuel ethanol from bamboo fibers | |
| CN101696427B (en) | Method for producing fuel ethanol and 2,3-butanediol by using fibrous matter | |
| BRPI0612966A2 (en) | Biomass treatment method | |
| BR122016004370B1 (en) | PROCESS FOR ETHANOL PREPARATION OF LIGNOCELLULOSTIC MATERIAL | |
| CN104498562A (en) | Method for pretreating agricultural waste biomass by adopting sodium hydroxide/urea/water system | |
| JP5711873B2 (en) | Simultaneous saccharification and fermentation of cellulosic materials | |
| CN110713939A (en) | Strain for degrading lignocellulose source inhibitor under extremely low pH condition and application | |
| CN104694587A (en) | Method for producing lactic acid from bagasse | |
| CN101092639A (en) | Method for degrading stalk of saccharified crops through biologic enzyme | |
| Gao et al. | Lignocellulolytic enzyme cocktail produced by plant endophytic Chaetomium globosum exhibits a capacity for high-efficient saccharification of raw rice straw | |
| CN111733197A (en) | Method for enzymatic hydrolysis and saccharification of plant fiber raw material | |
| CN116814709A (en) | Method for producing lactic acid by pretreatment of lignocellulose raw material by hydrothermal auxiliary enzyme method | |
| BR112021012936A2 (en) | METHOD OF TREATMENT OF A LIGNOCELLULOSIC BIOMASS | |
| Buyukoztekin et al. | Enzymatic hydrolysis of organosolv-pretreated corncob and succinic acid production by Actinobacillus succinogenes | |
| CN106032542B (en) | Method for producing ethanol by fermenting cellulose hydrolysate | |
| CN101289677B (en) | Process for preparing ethanol by using cellulose-containing raw material | |
| Wang et al. | Optimal tartaric acid pretreatment of reed for bioethanol production by fed batch semi-synchronous saccharification fermentation | |
| CN111218490A (en) | Method for pretreating lignocellulose by using ammonia and auxiliary agent | |
| AU2021104292A4 (en) | Processing method for enzymatic hydrolysis and saccharification of waste straw cellulose | |
| CN101509018B (en) | Method for preparing ethanol by raw materials containing cellulose | |
| Erliana et al. | The effects of various pH and temperature to enhance lactic acid production using Lactobacillus casei and Lactobacillus rhamnosus |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| RJ01 | Rejection of invention patent application after publication |
Application publication date: 20201002 |
|
| RJ01 | Rejection of invention patent application after publication |