CN115725672A - Pretreatment method for improving enzymatic saccharification effect of shell biomass and application thereof - Google Patents

Pretreatment method for improving enzymatic saccharification effect of shell biomass and application thereof Download PDF

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CN115725672A
CN115725672A CN202211530267.2A CN202211530267A CN115725672A CN 115725672 A CN115725672 A CN 115725672A CN 202211530267 A CN202211530267 A CN 202211530267A CN 115725672 A CN115725672 A CN 115725672A
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shell
pretreatment
biomass
enzymatic saccharification
improving
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王永华
王俊鸿
蓝东明
王方华
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South China University of Technology SCUT
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Abstract

The invention belongs to the field of comprehensive utilization of agriculture and forestry biomass, and discloses a pretreatment method for improving enzymatic saccharification effect of shell biomass and application thereof. The method comprises the steps of pretreating extracted shell biomass by using an ethanolamine aqueous solution under a heating condition, then carrying out solid-liquid separation on a pretreated mixture, washing a solid part with a washing solution to obtain pretreated residues, recovering the ethanolamine solution from a liquid part through vacuum rotary evaporation for reuse, and introducing CO into a concentrated liquid part 2 The lignin is precipitated. The method can selectively remove lignin and retain cellulose and hemicellulose by simple and mild pretreatment under the conditions of no solvent synthesis step and no catalyst, obviously improves the enzymolysis saccharification effect, can recycle the ethanolamine solution, has low energy consumption in the whole process, low cost, simplicity, practicability, greenness and environmental protection, and has the advantages of low energy consumption in the fields of fruit shell biomass pretreatment and biofuel preparationHas good application prospect.

Description

Pretreatment method for improving enzymatic saccharification effect of shell biomass and application thereof
Technical Field
The invention belongs to the field of comprehensive utilization of agriculture and forestry biomass, and particularly relates to a pretreatment method for improving enzymatic saccharification effect of shell biomass and application thereof.
Background
With the increasing shortage of fossil energy and the increasing demand for energy supply, the search and development of renewable energy sources are at hand. The shell biomass is one of lignocellulose biomass, belongs to agricultural and forestry waste, contains rich cellulose and hemicellulose, and can be hydrolyzed into fermentable monosaccharide so as to be further converted into biofuel. A large number of hydrogen bonds, covalent bonds and non-covalent bonds exist among cellulose, hemicellulose and lignin in biomass, a compact structural network is formed, the direct utilization of the biomass is prevented, pretreatment is usually needed to destroy the structure of the biomass, the solution permeability of the biomass raw material and the accessibility of enzyme are improved, and the biomass raw material is further promoted to be hydrolyzed and converted into monosaccharide for further utilization. Compared with straw agricultural and forestry wastes, the shell biomass usually contains more lignin, has harder texture and has higher pretreatment difficulty.
The biomass pretreatment method comprises a physical method, a chemical method, a biological method and a combined treatment method, wherein the physical method has high energy consumption, the biological method has long treatment period, and the chemical method or the combined use of the chemical method and other methods is the mainstream biomass pretreatment method at present. Among them, the eutectic solvent (DES) is a new generation of Ionic Liquid (IL), generally considered to have the advantages of no toxicity, good biocompatibility, low cost, environmental protection, etc.
In the prior art, a patent granted document with an authorization publication number of CN 108441530B discloses a method for pretreating straw lignocellulose by using an alkaline eutectic solvent, so that the content of hemicellulose sugar in pretreated residues and the total sugar yield after enzymolysis are improved, wherein the alcamines eutectic solvent has better performance. However, the eutectic solvent is generally synthesized by heating 2 or more organic solvents, which increases the pretreatment cost, and the introduction of water during the pretreatment process affects the pretreatment effect and the recovery of the subsequent solvent, thereby easily causing environmental pollution.
Qammer et al (Qammer Z., matthew J.E., et al. Arc de electric solutions return green. Therefore, the development of an economic, efficient, environment-friendly and sustainable fruit shell biomass pretreatment method is the key point for promoting waste utilization and large-scale production of biological energy.
Disclosure of Invention
In view of the above disadvantages of the prior art, the primary object of the present invention is to provide a pretreatment method for improving the enzymatic saccharification effect of shell biomass. The method utilizes alkaline ethanolamine solution, can selectively remove lignin, retain cellulose and most hemicellulose, effectively improve enzymolysis efficiency of pretreated residues, and increase total sugar yield. The method has the advantages of low cost, easy operation, recyclable solvent and environmental protection.
It is a further object of the present invention to provide the use of the above process in the production of biofuels.
The purpose of the invention is realized by the following technical scheme:
a pretreatment method for improving enzymatic saccharification effect of shell biomass comprises the following steps:
(1) Crushing a shell biomass raw material, extracting the crushed shell biomass raw material to be colorless by using an acetone aqueous solution, and drying the extracted raw material;
(2) Mixing the raw material extracted in the step (1) and an ethanolamine aqueous solution according to a certain solid-to-liquid ratio, and carrying out heating pretreatment;
(3) Separating the solid-liquid mixture pretreated in the step (2) to obtain a solid component and a liquid component, and washing the solid component with a washing liquid to obtain pretreated residues rich in cellulose and hemicellulose;
(4) Performing rotary evaporation on the liquid component in the step (3), recovering ethanolamine aqueous solution, and introducing CO into the concentrated liquid component 2 Precipitating lignin;
(5) And (4) adding the pretreated residues in the step (3) into an enzymolysis system consisting of compound cellulase, sodium citrate buffer solution and water according to a certain mass concentration for enzymolysis to obtain fermentable sugar.
Further, the fruit shell biomass raw material in the step (1) is one or more than two of oil tea fruit shell, coconut shell, olive shell, oil palm shell, walnut shell or almond shell; more preferably, the biomass raw material is agricultural and forestry waste camellia oleifera shells.
Further, the solid-to-liquid ratio of the raw material to the ethanolamine solution in the step (2) is 1; more preferably, the solid-to-liquid ratio of the raw material to the ethanolamine solution is 1.
Further, the concentration of the ethanolamine aqueous solution in the step (2) is 25-100% (w/w); more preferably, the concentration of the ethanolamine aqueous solution is 75% to 100%.
Further, the concentration of the acetone aqueous solution is 50-90% (v/v), and the pretreatment conditions in the step (2) are as follows: the temperature is 70-130 ℃, the time is 1-3h, and the stirring speed is 500-1000rpm; more preferably, the concentration of the acetone aqueous solution is 80%, the temperature is 90-130 ℃, the time is 1-3h, and the stirring speed is 500rpm.
Further, the pulverization in the step (1) is to be pulverized to 40-60 meshes, and the drying is hot air drying; the solid-liquid separation mode in the step (3) is centrifugation or vacuum filtration; more preferably, the solid-liquid separation method is centrifugal separation.
Further, the washing solution in the step (3) is ethanol or water; more preferably, the washing solution is deionized water.
Further, CO described in the step (4) 2 The introduction conditions are as follows: the pressure is 0.2-0.5MPa, and the time is 10-30min; more preferably, said CO 2 The introduction conditions are as follows: the pressure is 0.2MPa and the time is 20min.
Further, the mass concentration of the pretreatment residue in the step (5) is 1% -5%, the composite cellulase is Novovern Celluclast 1.5L, cellicCTec 2 or CellicCTec 3, and the addition amount is 20-100FPU/g pretreatment residue; more preferably, the mass concentration of the pretreatment residue is 1%, the composite cellulase is Novoxil CellicCTec 3, and the addition amount is 20FPU/g pretreatment residue.
Further, the enzymolysis conditions in the step (5) are as follows: the pH is 4.8-5.5, the temperature is 45-55 ℃, and the enzymolysis time is 48-96h; more preferably, the enzymolysis conditions are as follows: the pH value is 5.0, the temperature is 50 ℃, and the enzymolysis time is 72h.
The use of the above process in the production of biofuels.
Compared with the prior art, the invention has the following advantages and beneficial effects:
(1) The pretreatment method provided by the invention is simple and easy to operate, low in cost, non-toxic and harmless, and environment-friendly, and the solvent can be recovered.
(2) The pretreatment method provided by the invention can selectively remove lignin, retain cellulose and most hemicellulose, remarkably improve the enzymolysis saccharification effect of the pretreatment residues, increase the yield of fermentable sugar which can reach 79.43 percent and is beneficial to later-stage conversion into biofuel.
(3) The pretreatment method of the shell biomass provided by the invention has good application potential in the field of biofuel preparation.
Drawings
FIG. 1 is a flow chart of the pretreatment method of the present invention.
Detailed Description
The present invention will be described in further detail with reference to specific examples, but the embodiments of the present invention are not limited thereto, and the process parameters not specifically mentioned may be performed with reference to conventional techniques.
The composite cellulase CellicCTec 3 is purchased from Novovin (China) investment Limited.
Example 1
A pretreatment method for improving enzymatic saccharification effect of shell biomass comprises the following steps:
pulverizing oil tea fruit shell raw material to 40-60 mesh, extracting with 80% acetone water solution (v/v), and oven drying at constant temperature of 60 deg.C. Weighing a certain amount of raw materials after extraction, adding 100% ethanolamine solution according to the solid-to-liquid ratio of 1. After the pretreatment is finished, performing solid-liquid separation in a centrifugal mode, washing the solid part to be colorless by using deionized water to obtain pretreatment residues, and freezing and storing the pretreatment residues for subsequent enzymolysis of sugarAnd (4) transforming. Rotary evaporating the liquid part, recovering ethanolamine, and introducing 0.2MPa CO into the concentrated liquid component 2 For 20min to precipitate lignin. Adding the pretreatment residues into a sodium citrate buffer solution with the pH value of 4.8 according to the mass concentration of 1%, adding a composite cellulase CellicCTec 3 according to the proportion of 20FPU/g of the pretreatment residues, and performing enzymolysis for 72 hours in a water bath shaker at 50 ℃ and 150rpm to obtain the fermentable sugar.
Example 2
A pretreatment method for improving enzymatic saccharification effect of shell biomass comprises the following steps:
pulverizing oil tea fruit shell raw material to 40-60 mesh, extracting with 80% acetone water solution (v/v), and oven drying at constant temperature of 60 deg.C. Weighing a certain amount of raw materials after extraction, adding 100% ethanolamine solution according to the solid-to-liquid ratio of 1. And after the pretreatment is finished, performing solid-liquid separation in a centrifugal mode, washing the solid part to be colorless by using deionized water to obtain pretreatment residues, and freezing and storing the pretreatment residues for subsequent enzymolysis and saccharification. Rotary evaporating the liquid part, recovering ethanolamine, and introducing 0.2MPa CO into the concentrated liquid component 2 For 20min to precipitate lignin. Adding the pretreated residues into a sodium citrate buffer solution with the pH value of 4.8 according to the mass concentration of 1%, adding the composite cellulase CellicCTec 3 according to the proportion of 20FPU/g of the pretreated residues, and performing enzymolysis for 72 hours in a water bath shaking table at 50 ℃ and 150rpm to obtain the fermentable sugar.
Example 3
A pretreatment method for improving enzymatic saccharification effect of shell biomass comprises the following steps:
pulverizing oil tea fruit shell raw material to 40-60 mesh, extracting with 80% acetone water solution (v/v), and oven drying at 60 deg.C. Weighing a certain amount of the raw materials after extraction, adding 100% ethanolamine solution according to the solid-to-liquid ratio of 1. And after the pretreatment is finished, performing solid-liquid separation in a centrifugal mode, washing the solid part to be colorless by using deionized water to obtain pretreatment residues, and freezing and storing the pretreatment residues for subsequent enzymolysis and saccharification. Rotary evaporating the liquid part, recovering ethanolamine, and introducing 0.2MP into the concentrated liquid componenta CO of 2 For 20min to precipitate lignin. Adding the pretreated residues into a sodium citrate buffer solution with the pH value of 4.8 according to the mass concentration of 1%, adding the composite cellulase CellicCTec 3 according to the proportion of 20FPU/g of the pretreated residues, and performing enzymolysis for 72 hours in a water bath shaking table at 50 ℃ and 150rpm to obtain the fermentable sugar.
Example 4
A pretreatment method for improving enzymatic saccharification effect of shell biomass comprises the following steps:
pulverizing oil tea fruit shell raw material to 40-60 mesh, extracting with 80% acetone water solution (v/v), and oven drying at constant temperature of 60 deg.C. Weighing a certain amount of the raw materials after extraction, adding 100% ethanolamine solution according to the solid-to-liquid ratio of 1. And after the pretreatment is finished, performing solid-liquid separation in a centrifugal mode, washing the solid part to be colorless by using deionized water to obtain pretreatment residues, and freezing and storing the pretreatment residues for subsequent enzymolysis and saccharification. Rotary evaporating the liquid part, recovering ethanolamine, and introducing 0.2MPa CO into the concentrated liquid component 2 For 20min to precipitate lignin. Adding the pretreatment residues into a sodium citrate buffer solution with the pH value of 4.8 according to the mass concentration of 1%, adding a composite cellulase CellicCTec 3 according to the proportion of 20FPU/g of the pretreatment residues, and performing enzymolysis for 72 hours in a water bath shaker at 50 ℃ and 150rpm to obtain the fermentable sugar.
Example 5
A pretreatment method for improving enzymatic saccharification effect of shell biomass comprises the following steps:
pulverizing oil tea fruit shell raw material to 40-60 mesh, extracting with 80% acetone water solution (v/v), and oven drying at 60 deg.C. Weighing a certain amount of the raw materials after extraction, adding 50% ethanolamine aqueous solution (w/w) according to the solid-to-liquid ratio of 1. And after the pretreatment is finished, performing solid-liquid separation in a centrifugal mode, washing the solid part to be colorless by using deionized water to obtain pretreatment residues, and freezing and storing the pretreatment residues for subsequent enzymolysis and saccharification. Rotary evaporating the liquid part, recovering ethanolamine, and introducing 0.2MPa CO into the concentrated liquid component 2 For 20min to precipitate lignin. The pretreatment residue is added according to the mass concentration of 1 percentAdding into sodium citrate buffer solution with pH of 4.8, adding composite cellulase CellicCTec 3 according to the proportion of 20FPU/g pretreatment residue, and performing enzymolysis for 72h in a water bath shaker at 50 deg.C and 150rpm to obtain fermentable sugar.
Example 6
After the ethanolamine aqueous solution finishes treating the oil-tea camellia shells, the recycling comprises the following steps:
in example 3, the ethanolamine solution is recovered by evaporating the liquid part in a vacuum rotary evaporator at 45 ℃, and the recovered ethanolamine solution is directly pretreated according to the steps described in example 3 without purification, and is recycled for 4 times.
Comparative example 1
In order to fully show that the enzymolysis saccharification effect of the oil-tea camellia shell pretreatment residues can be effectively improved by ethanolamine aqueous solution treatment, and the monosaccharide yield of direct enzymolysis of the oil-tea camellia shell raw materials is analyzed, as a comparative example, the method comprises the following steps:
pulverizing oil tea fruit shell raw material to 40-60 mesh, extracting with 80% acetone water solution (v/v), and oven drying at constant temperature of 60 deg.C. Adding the extracted oil tea fruit shell into a sodium citrate buffer solution with the pH value of 4.8 according to the mass concentration of 1%, adding composite cellulase CellicCTec 3 according to the proportion of 20FPU/g oil tea fruit shell raw material, and carrying out enzymolysis for 72h in a water bath shaking table at 50 ℃ and 150rpm to obtain the fermentable sugar.
Comparative example 2
In order to further illustrate that the pretreatment of the ethanolamine aqueous solution improves the enzymolysis saccharification effect and reduces the pretreatment cost, the choline chloride-ethanolamine eutectic solvent with the molar ratio of 1:
pulverizing oil tea fruit shell raw material to 40-60 mesh, extracting with 80% acetone water solution (v/v), and oven drying at constant temperature of 60 deg.C. Mixing two reagents of choline chloride and ethanolamine according to a molar ratio of 1. Weighing a certain amount of raw materials after extraction, adding a choline chloride-ethanolamine eutectic solvent according to a solid-liquid ratio of 1. And after the pretreatment is finished, performing solid-liquid separation in a centrifugal mode, washing the solid part with deionized water until the solid part is colorless to obtain pretreatment residues, and freezing and storing the pretreatment residues for subsequent enzymolysis and saccharification. Adding the pretreatment residues into a sodium citrate buffer solution with the pH value of 4.8 according to the mass concentration of 1%, adding a composite cellulase CellicCTec 3 according to the proportion of 20FPU/g of the pretreatment residues, and performing enzymolysis for 72 hours in a water bath shaker at 50 ℃ and 150rpm to obtain the fermentable sugar.
Test example
The components of the pretreated camellia oleifera shells are analyzed according to the National Renewable Energy Laboratory (NREL) standard analysis method, and the monosaccharide content in the enzymatic hydrolysate is determined by adopting high performance liquid chromatography.
As can be seen from the data in Table 1, the pretreatment method of the husk biomass in the embodiment of the invention can selectively remove lignin, almost completely retain cellulose and most polyxylose, has obvious deconstruction effect, and has the glucose and xylose yields 2.75-4.24 times and 49.96-70.79 times of the untreated biomass respectively after enzymolysis. Meanwhile, after the ethanolamine solution is recycled for four times, the yields of glucose and xylose after the enzymolysis of the pretreated residues still reach 83.86 percent and 71.70 percent respectively, and the recoverability is good. In addition, under the same condition, compared with the deep eutectic solvent pretreatment formed by heating ethanolamine and choline chloride, the ethanolamine solution pretreatment can obtain the same or higher enzymolysis sugar yield, and further proves that the method has the characteristics of low cost, simplicity and easiness in operation, sustainability and the like, and has wide application potential in the field of biomass fuels.
Table 1: component change and enzymolysis sugar yield (%) -before and after pretreatment of oil tea fruit shell
Figure BDA0003974521710000091
Note: solids recovery refers to the ratio of the weight of solids remaining after pretreatment to the weight of solid biomass before pretreatment.
The above embodiments are only preferred embodiments of the present invention, but the embodiments of the present invention are not limited by the above embodiments, and any other changes, modifications, substitutions, combinations, and simplifications which do not depart from the spirit and principle of the present invention should be regarded as equivalent replacements within the protection scope of the present invention.

Claims (10)

1. A pretreatment method for improving the enzymatic saccharification effect of shell biomass is characterized by comprising the following steps:
(1) Crushing a shell biomass raw material, extracting the crushed shell biomass raw material to be colorless by using an acetone aqueous solution, and drying the extracted raw material;
(2) Mixing the raw material extracted in the step (1) and an ethanolamine aqueous solution according to a certain solid-to-liquid ratio, and carrying out heating pretreatment;
(3) Separating the solid-liquid mixture pretreated in the step (2) to obtain a solid component and a liquid component, and washing the solid component with a washing solution to obtain pretreatment residues rich in cellulose and hemicellulose;
(4) Performing rotary evaporation on the liquid component in the step (3), recovering ethanolamine aqueous solution, and introducing CO into the concentrated liquid component 2 Precipitating lignin;
(5) And (4) adding the pretreated residues in the step (3) into an enzymolysis system consisting of compound cellulase, sodium citrate buffer solution and water according to a certain mass concentration for enzymolysis to obtain the fermentable sugar.
2. The pretreatment method for improving enzymatic saccharification effect of shell-type biomass as claimed in claim 1, wherein in step (1), the shell-type biomass raw material is one or more than two of oil tea fruit shell, coconut shell, olive shell, oil palm shell, walnut shell or almond shell.
3. The pretreatment method for improving the enzymatic saccharification effect of the shell biomass according to claim 1, wherein the solid-liquid ratio of the raw material to the ethanolamine solution in the step (2) is 1.
4. The pretreatment method for improving enzymatic saccharification effect of fruit shell biomass according to claim 1, wherein the concentration of the ethanolamine aqueous solution in the step (2) is 25% -100%.
5. The pretreatment method for improving the enzymatic saccharification effect of the shell biomass as claimed in claim 1, wherein the concentration of the acetone aqueous solution in the step (1) is 50% -90%, and the pretreatment conditions in the step (2) are as follows: the temperature is 70-130 ℃, the time is 1-3h, and the stirring speed is 500-1000rpm.
6. The pretreatment method for improving enzymatic saccharification effect of fruit shell biomass according to claim 1, wherein the crushing in step (1) is crushing to 40-60 meshes, and the drying is hot air drying;
the solid-liquid separation mode in the step (3) is centrifugation or vacuum filtration, and the washing liquid in the step (3) is ethanol or water.
7. The pretreatment method for improving enzymatic saccharification effect of fruit shell biomass as claimed in claim 1, wherein the CO in step (4) 2 The introduction conditions are as follows: the pressure is 0.2-0.5MPa, and the time is 10-30min.
8. The pretreatment method for improving the enzymatic saccharification effect of the shell-type biomass as claimed in claim 1, wherein the mass concentration of the pretreatment residues in the step (5) is 1% -15%, the composite cellulase is Novexin Celluclast 1.5L, cellicCTec 2 or CellicCTec 3, and the addition amount is 20-50FPU/g of pretreatment residues.
9. The pretreatment method for improving enzymatic saccharification effect of fruit shell biomass according to claim 1, wherein the enzymatic hydrolysis conditions in step (5) are as follows: the pH is 4.8-5.5, the temperature is 45-55 ℃, and the enzymolysis time is 24-72h.
10. Use of a process according to any one of claims 1 to 9 in the production of a biofuel.
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