High-yield enzymolysis saccharification method for corncob acid-alkali step-by-step impurity removal

Abstract

The invention relates to the technical field of biomass energy, and discloses a corncob acid-alkali step-by-step impurity removal high-yield enzymolysis saccharification method which comprises the following specific steps: preparing raw materials; carrying out acid and alkali two-step pretreatment; collecting the pretreated corncobs, and repeatedly washing the corncobs for multiple times until the corncobs are neutral to obtain washed corncobs; according to the method, the corncobs are pretreated in the two steps of acid and alkali, so that part of hemicellulose and lignin in the corncobs are removed, the porosity of the corncobs is increased, cellulose is moistened and expanded, the enzyme hydrolysis yield is increased, and the yield of glucose production is increased. According to the method, sodium hydroxide and calcium hydroxide are selected for combined pretreatment of the corncobs, so that the cost is lower, SO4 < 2 + > generated by acid treatment can neutralize Ca < 2 + > generated by alkali treatment, the cost is low, the environmental pollution is less, a corncob saccharification process with lower cost is provided, the enzymatic hydrolysate can be further fermented and converted into lactic acid and ethanol products subsequently by utilizing microorganisms, and the method is suitable for industrial production. The economic value is improved.

Description

Technical Field

[0001] The present invention relates to the technical field of biomass energy, and particularly to a high-yield enzymatic hydrolysis saccharification method for stepwise removing impurities from corncobs by acid and alkali. Background Art

[0002] With the continuous development of the economy, people's demand for and consumption of energy have been increasing year by year, and the energy crisis has become one of the major challenges faced by human society. Compared with fossil energy, biomass resources (especially lignocellulose) as the most abundant renewable resources in nature have the advantages of wide sources, renewable resources, and environmental friendliness, and are expected to replace fossil energy and become an important direction for the development of new energy.

[0003] The preparation of fuel ethanol from cellulosic biomass is of great significance for new energy. This technology includes steps such as raw material pretreatment, hydrolysis saccharification, and ethanol fermentation. Among them, how to efficiently convert cellulose raw materials into fermentable reducing sugars has become the key point of research. However, so far in current research, due to the influence of various factors on cellulose enzymatic hydrolysis, the efficiency of cellulose enzymatic hydrolysis is low and the glucose yield is low. Therefore, finding a method for efficient enzymatic hydrolysis of cellulose is of great significance.

[0004] The main factors affecting cellulose enzymatic hydrolysis include raw material pretreatment, reaction temperature, pH value, enzyme dosage, substrate mass concentration, etc. Among them, the purpose of raw material pretreatment is to reduce the recalcitrant characteristics of lignocellulose (including the chemical characteristics of lignin and hemicellulose), thereby promoting the enzymatic hydrolysis process (increasing the accessibility of enzymes). Chemical pretreatment is one of the pretreatment methods, including acid treatment, alkali treatment, organic solvent treatment, and ionic liquid treatment. However, this single pretreatment method often has disadvantages such as harsh treatment conditions, high treatment costs, and low enzymatic hydrolysis efficiency, which limit its industrial production.

[0005] The two-step acid and alkali pretreatment method can stepwise remove hemicellulose and lignin from the raw material, enabling the hierarchical separation of cellulose, hemicellulose, and lignin in the raw material, providing the possibility for the distributed utilization of the three components. Cellulose is well enriched in the material, which is very beneficial for further enzymatic hydrolysis saccharification and lactic acid fermentation.

[0006] China is a large agricultural country with a large amount of agricultural and forestry waste. For example, the annual output of corncobs is about 30 million tons. Corncobs are a typical lignocellulosic biomass, mainly composed of cellulose, hemicellulose, lignin, etc. At present, most corncobs are burned as rural fuels, which is not only a waste of resources but also causes a series of environmental pollution problems. Using corncobs as raw materials for enzymatic hydrolysis saccharification can not only comprehensively utilize agricultural product resources, protect the environment, but also increase farmers' income, produce biofuels and high-value-added chemicals, and has good economic and social benefits. Summary of the Invention

[0007] (1) Technical problems to be solved

[0008] In view of the deficiencies of the prior art, the present invention provides a high-yield enzymatic hydrolysis and saccharification method for stepwise acid-alkali impurity removal from corncobs, which has the advantages of high-efficiency enzymatic hydrolysis, high glucose yield, environmental friendliness and controllable cost, and solves the problems of low cellulase hydrolysis efficiency, low glucose yield, limited industrialization of single pretreatment methods, waste of corncob resources and environmental pollution.

[0009] (2) Technical solutions

[0010] To achieve the above object, the present invention provides the following technical solutions: A high-yield enzymatic hydrolysis and saccharification method for stepwise acid-alkali impurity removal from corncobs, comprising the following steps:

[0011] S1. Raw material preparation: Air-dry, crush and screen corncobs to obtain treated corncobs, and put the obtained treated corncobs into a sealed bag for subsequent use;

[0012] S2. Acid pretreatment: Weigh the treated corncobs of step S1 in a formula ratio, put them into a conical flask and place it in a water bath kettle, and add sulfuric acid for reaction under the conditions of controlling the solid-liquid ratio of the raw material suspension, sulfuric acid concentration and the temperature of the water bath kettle;

[0013] S3. Post-treatment after acid treatment: After the reaction reaches 30 - 120 min, collect the reacted corncobs, wash the corncobs until neutral, collect the washing liquid, and dry and store the reacted corncobs;

[0014] S4. Alkali pretreatment: Weigh the corncobs after acid treatment of step S3, put them into a conical flask and place it in a water bath kettle, and add sodium hydroxide and calcium hydroxide for reaction under the conditions of controlling the solid-liquid ratio of the raw material suspension, the ratio of sodium hydroxide to calcium hydroxide, the total addition amount of sodium hydroxide and calcium hydroxide in the raw material suspension and the temperature of the water bath kettle;

[0015] S5. Post-treatment after alkali treatment: After the reaction reaches 1 h - 5 h, collect the reacted corncobs, wash the corncobs until neutral, collect the washing liquid, and dry and store the reacted corncobs;

[0016] S6. Treatment of treatment liquid: After the collected acid treatment liquid and alkali treatment liquid are comprehensively utilized subsequently, neutralize and discharge;

[0017] S7. Enzymatic hydrolysis: Weigh the corncobs after two-step acid and alkali pretreatment as the enzymatic hydrolysis substrate, put them into a conical flask and place it in a magnetic stirring water bath kettle, and add cellulase for enzymatic hydrolysis under the conditions of controlling the substrate concentration, cellulase dosage, pH range and constant temperature stirring conditions to obtain a hydrolysis solution, that is, complete the high-efficiency enzymatic hydrolysis of corncobs pretreated by two-step acid and alkali to produce glucose.

[0018] Preferably, in the step S1, the components of the corncob raw material are: cellulose content 34% - 38%, hemicellulose content 20% - 30%, lignin and other contents 30% - 40%.

[0019] Preferably, in the step S1, the screening is carried out with a 30 - mesh sieve.

[0020] Preferably, in the S2, the conditions for acid pretreatment are: controlling the solid - liquid ratio of the raw material suspension at 1:16 - 1:10, controlling the sulfuric acid concentration of the raw material suspension at 0.7% - 2.2%, and controlling the temperature of the water bath at 60 - 90°C.

[0021] Preferably, the post - treatment process after acid treatment in the S3 is as follows:

[0022] S3.1. After the reaction reaches 30 - 120 min, collect the reacted corncobs and wash them repeatedly several times until the corncobs are neutral, and collect the washing liquid;

[0023] S3.2. Place the reacted corncobs in an oven at 105°C to dry, and obtain the corncobs after acid treatment;

[0024] S3.3. Store it in an environment at 2 - 5°C for subsequent use.

[0025] Preferably, the conditions for alkali pretreatment in the S4 are: controlling the solid - liquid ratio of the raw material suspension at 1:16 - 1:10, controlling the ratio of sodium hydroxide to calcium hydroxide between 3:1 - 6:1, controlling the total addition amount of sodium hydroxide and calcium hydroxide in the raw material suspension at 0.03 - 0.15 g / g of the substrate, and controlling the temperature of the water bath at 30 - 60°C.

[0026] Preferably, the post - treatment process after alkali treatment in the S5 is as follows:

[0027] S5.1. After the reaction reaches 1 h - 5 h, collect the reacted corncobs and wash them repeatedly several times until neutral, and collect the washing liquid;

[0028] S5.2. Place the reacted corncobs in an oven at 105°C to dry, and obtain the corncobs after two - step pretreatment with acid and alkali;

[0029] S5.3. Store it in a low - temperature environment at 2 - 5°C for subsequent use.

[0030] Preferably, the specific process of enzymatic hydrolysis in the S7 is as follows:

[0031] S7.1. Weigh the corncobs after two - step pretreatment with acid and alkali as the substrate for enzymatic hydrolysis, put them into a conical flask and place it in a magnetic stirring water bath.

[0032] S7.2. Add distilled water to control the substrate concentration at 4% - 12%, and stir magnetically to evenly disperse the raw materials.

[0033] S7.3. Then add cellulase into the conical flask so that the dosage of cellulase in the whole hydrolysis system is 0.01 - 0.05 g / g of substrate, and control the pH range at 4.0 - 6.0.

[0034] S7.4. Finally, seal the bottle mouth with plastic wrap, place it under constant temperature conditions and stir continuously for enzymatic hydrolysis to obtain the hydrolysis solution, thus completing the high-efficiency enzymatic hydrolysis of corncob pretreated by acid and alkali steps to produce glucose.

[0035] Preferably, the conditions for enzymatic hydrolysis treatment in S7 are: the hydrolysis temperature is 40 - 60 °C, the hydrolysis time is 24 - 72 h, and the rotation speed is 400 - 500 rpm.

[0036] Preferably, the pH range is controlled between 4.0 - 6.0 in step S7, and the acids and alkalis used to control the pH are hydrochloric acid solution and sodium hydroxide solution.

[0037] Compared with the prior art, the present invention provides a high-yield enzymatic saccharification method for stepwise impurity removal of corncob by acid and alkali, having the following beneficial effects:

[0038] 1. By pretreating corncob by acid - alkali steps, the present invention can effectively remove the impurities of corncob, improve the accessibility of cellulose and hemicellulose in corncob, create favorable conditions for subsequent enzymatic hydrolysis, and thus increase the glucose yield in corncob. During the acid treatment process, part of the hemicellulose and lignin in corncob degrade, initially destroying its internal complex structure; while the subsequent alkali treatment can further remove lignin and further destroy the structure of corncob, making cellulose more easily acted upon by enzymes. Through this two-step acid and alkali pretreatment method, under optimized acid and alkali pretreatment conditions, the tightly bound structure of cellulose, hemicellulose and lignin in corncob can be effectively destroyed, and the crystallinity of cellulose can be reduced, thus achieving an efficient enzymatic hydrolysis effect. The above treatment method can greatly improve the hydrolysis conversion rate of corncob and provide a more efficient raw material basis for subsequent biological conversion processes.

[0039] 2. By precisely controlling various parameters in the enzymatic hydrolysis process, such as substrate concentration, cellulase dosage, pH range, hydrolysis temperature and time, the present invention achieves the beneficial effects of optimizing the enzymatic hydrolysis reaction, improving the enzymatic saccharification efficiency, and increasing the glucose concentration in the hydrolysis solution. The combination of different parameters has an obvious influence on the enzymatic hydrolysis effect, enabling the enzyme to fully play its role and efficiently converting cellulose into glucose.

[0040] 3. By comprehensively utilizing the acid treatment liquid and the alkali treatment liquid, the present invention achieves the beneficial effect of reasonable disposal of the treatment liquid. After neutralization treatment and discharge, it can not only reduce environmental pollution, but also extract valuable components from the treatment liquid to realize the recycling of resources, thereby reducing production costs and improving process economy. At the same time, the raw material of the present invention is agricultural waste - corncob. Extracting glucose from corncob can effectively reduce the input cost of biological enzyme hydrolysis, reduce the environmental pollution load, relieve the demand for fossil energy, and ultimately reduce the usage amount of lignocellulosic raw materials, thus conforming to the concept of green and sustainable development. Therefore, under the premise of reducing the input cost of enzyme hydrolysis, the method of the present invention achieves the purpose of high-efficiency hydrolysis and saccharification of corncob, not only with a high hydrolysis yield, but also with economic benefits. BRIEF DESCRIPTION OF THE DRAWINGS

[0041] Figure 1 It is a process step diagram of the method of the present invention. DETAILED DESCRIPTION OF THE EMBODIMENTS

[0042] Next, the technical solutions in the embodiments of the present invention will be clearly and completely described in conjunction with the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative efforts shall fall within the protection scope of the present invention.

[0043] Please refer to Figure 1 , a high-yield enzymatic hydrolysis and saccharification method for stepwise acid-alkali impurity removal of corncob, comprising the following steps:

[0044] S1. Raw material preparation: Air-dry, crush, and screen the corncob to obtain the treated corncob, and put the obtained treated corncob into a sealed bag for subsequent use;

[0045] S2. Acid pretreatment: Weigh the treated corncob in step S1 according to the formulated ratio, put it into a conical flask and place it in a water bath kettle, and add sulfuric acid (sulfuric acid) to react under the conditions of controlling the solid-liquid ratio of the raw material suspension, the sulfuric acid concentration, and the temperature of the water bath kettle;

[0046] S3. Post-treatment after acid treatment: After the reaction reaches 30 - 120 min, collect the reacted corncob, wash the corncob until it is neutral, collect the washing liquid, and dry and store the reacted corncob;

[0047] S4. Alkali pretreatment: Weigh the acid-treated corncob in step S3, put it into a conical flask and place it in a water bath kettle, and add sodium hydroxide and calcium hydroxide to react under the conditions of controlling the solid-liquid ratio of the raw material suspension, the ratio of sodium hydroxide to calcium hydroxide, the total addition amount of sodium hydroxide and calcium hydroxide in the raw material suspension, and the temperature of the water bath kettle;

[0048] S5. Post-treatment with alkali: After the reaction reaches 1 h - 5 h, collect the corncobs after the reaction, wash the corncobs until neutral, collect the washing liquid, and dry and store the corncobs after the reaction.

[0049] Advantages: By pretreating corncobs step by step with acid and alkali, impurities in the corncobs can be effectively removed, the accessibility of cellulose and hemicellulose in the corncobs can be improved, creating favorable conditions for subsequent enzymatic hydrolysis, and thus the glucose yield in the corncobs can be increased. During the acid treatment process, part of the hemicellulose and lignin in the corncobs are degraded, initially destroying their internal complex structure; while the subsequent alkali treatment can further remove lignin and further destroy the structure of the corncobs, making cellulose more easily acted upon by enzymes. Through this two-step acid and alkali pretreatment method, under optimized acid and alkali pretreatment conditions, the tightly bound structure of cellulose, hemicellulose, and lignin in the corncobs can be effectively destroyed, and the crystallinity of cellulose can be reduced, thereby achieving an efficient enzymatic hydrolysis effect. The above treatment method can greatly improve the hydrolysis conversion rate of corncobs, providing a more efficient raw material basis for subsequent biological conversion processes.

[0050] S6. Treatment of the treatment liquid: After the collected acid treatment liquid and alkali treatment liquid are comprehensively utilized later, they are neutralized and discharged.

[0051] S7. Enzymatic hydrolysis: Weigh the corncobs pretreated by the two-step acid and alkali method as the substrate for enzymatic hydrolysis, put them into a conical flask and place it in a magnetic stirring water bath. Under the conditions of controlling the substrate concentration, the dosage of cellulase, the pH range, and constant temperature stirring, add cellulase for enzymatic hydrolysis to obtain a hydrolysis liquid, that is, the high-efficiency enzymatic hydrolysis of corncobs pretreated by the two-step acid and alkali method to produce glucose is completed.

[0052] Advantages: The present invention realizes the beneficial effect of reasonable disposal of the treatment liquid by comprehensively utilizing the acid treatment liquid and alkali treatment liquid. After neutralization treatment and then discharge, it can not only reduce environmental pollution, but also extract valuable components from the treatment liquid to realize resource recycling, thereby reducing production costs and improving process economy. At the same time, the raw material of the present invention is agricultural waste - corncobs. Extracting glucose from corncobs can effectively reduce the input cost of biological enzymatic hydrolysis, reduce the environmental pollution load, relieve the demand for fossil energy, and finally reduce the usage amount of lignocellulosic raw materials, thus conforming to the concept of green and sustainable development.

[0053] Compared with the prior art, a method for efficiently hydrolyzing corncob to produce glucose by two-step pretreatment with acid and alkali according to the present invention uses agricultural waste corncob as a raw material. First, the corncob is acid-treated to remove hemicellulose in the corncob, and then alkali-treated to remove lignin in the corncob, swell the cellulose in the corncob, and improve the accessibility of cellulase, thereby increasing the hydrolysis rate of the corncob. In addition, by comprehensively using SO4 2+ produced by acid treatment and Ca 2+ produced by alkali treatment, the pollutant emissions and treatment costs can be significantly reduced. This method can fractionate cellulose, hemicellulose, and lignin in the corncob, increase the hydrolysis rate of cellulose, provide a basis for subsequent production of high-value products, and provide certain technical support for the high-value utilization of corncob cellulose.

[0054] The corncob is treated by a two-step pretreatment method with acid and alkali to calculate the hemicellulose removal rate and lignin removal rate. Specifically:

[0055] Cellulose determination: Weigh 0.05 g of corncob raw material, add 5 mL of a mixed solution of acetic acid and nitric acid with a volume ratio of 2:1, shake well, and place in a boiling water bath for 25 min. After cooling, centrifuge and discard the supernatant, wash with distilled water by centrifugation 3 times. Then add 10 mL of 10% sulfuric acid and 10 mL of 0.1 mol / L potassium dichromate solution to the precipitate, shake well, and place in a boiling water bath for 10 min. After taking out, pour it into a conical flask. After the solution cools, add 5 mL of 20% potassium iodide solution and 1 mL of 0.5% starch solution, and titrate with 0.2 mol / L sodium thiosulfate solution until the solution turns dark green. In addition, titrate 10 mL of 10% sulfuric acid and 10 mL of 0.1 mol / L potassium dichromate solution as a blank control. The cellulose content is calculated according to formula (1): 10n

[0056]

[0057] In the formula, a is the volume of sodium thiosulfate consumed in the blank titration, in ml; b is the volume of sodium thiosulfate solution consumed by the solution, in ml; n is the mass of the corncob, in g; 0.9 is the coefficient for converting hexose to hemicellulose; 248 is the molecular weight of sodium thiosulfate.

[0058] Determination of hemicellulose: Weigh 0.05 g of corncob raw material, add 15 mL of 80% calcium nitrate solution, heat to boiling for 5 min, cool, and centrifuge at 5000 r / min for 5 min. Wash the precipitate 3 times with hot distilled water. Add 10 mL of 2 mol / L hydrochloric acid to the precipitate, place it in a boiling water bath for 45 min, cool, and centrifuge. Transfer the supernatant to a 100 mL volumetric flask. Rinse the precipitate 3 times with 10 mL of distilled water, combine the washing solutions in the volumetric flask, add 1 drop of phenolphthalein, neutralize to orange-red with 2 mol / L sodium hydroxide solution, dilute to the mark. Pipette 10 mL of the filtrate into a test tube, add 10 mL of alkaline copper reagent, boil in water for 15 min, cool, add 2 mL of oxalic acid and 0.5 mL of concentrated sulfuric acid, add 0.5 mL of 0.5% starch solution, and titrate with 0.01 mol / L sodium thiosulfate solution until the blue color disappears. Pipette another 10 mL of distilled water into a conical flask, add 10 mL of alkaline copper reagent, 2 mL of oxalic acid, 0.5 mL of concentrated sulfuric acid, and 0.5 mL of 0.5% starch, mix well, and titrate with 0.01 mol / L sodium thiosulfate solution until the blue color disappears as a blank control. The determination of hemicellulose content is calculated according to formula (2):

[0059]

[0060] In formula (3), 48 is the equivalent number of sodium thiosulfate corresponding to 1 mol of C6H10O5; K is the concentration of the sodium thiosulfate solution, with the unit of mol / L; a is the volume of the sodium thiosulfate solution consumed in the blank titration, in ml; b is the volume of the sodium thiosulfate solution consumed by the solution, with the unit of ml; n is the mass of the corncob, with the unit of g.

[0061] The following Examples 1-6 were obtained from the preparation process of the present invention:

[0062] Example 1

[0063] (1) Raw material preparation: Air-dry, crush, and screen the corncob to obtain the treated corncob, and put the obtained treated corncob into a sealed bag for subsequent use;

[0064] (2) Two-step acid and alkali pretreatment: Weigh the treated corncob in step (1), put it into a conical flask and place it in a water bath pot, add sulfuric acid, control the solid-liquid ratio of the raw material suspension to be 1:16, control the sulfuric acid concentration of the raw material suspension to be 2.2%, and control the temperature of the water bath pot to be 90 °C;

[0065] (3) After the reaction reaches 120 min, collect the reacted corncob and wash it repeatedly several times until it is neutral. Collect the washing solution, and dry the reacted corncob in an oven at 105 °C to obtain the acid-treated corncob, and store it in an environment of 2-5 °C for subsequent use;

[0066] (4) Weigh the acid-treated corncob in step (3), put it into a conical flask and place it in a water bath. Add sodium hydroxide and calcium hydroxide, control the solid-liquid ratio of the raw material suspension to be 1:16, control the ratio of sodium hydroxide to calcium hydroxide to be between 5:1, control the total addition amount of sodium hydroxide and calcium hydroxide in the raw material suspension to be 0.09 g / g of the substrate, and control the temperature of the water bath to be 60 °C;

[0067] (5) After the reaction reaches 3 h, collect the reacted corncob and wash it repeatedly several times until it is neutral. Collect the washing liquid, and place the reacted corncob in an oven at 105 °C to dry, obtaining the corncob pretreated by acid and alkali steps. Store it in a low-temperature environment of 2-5 °C for subsequent use;

[0068] (6) After the collected acid treatment liquid and alkali treatment liquid are comprehensively utilized later, neutralize and discharge them;

[0069] (7) Enzymatic hydrolysis: Weigh the corncob pretreated by acid and alkali steps as the enzymatic hydrolysis substrate, put it into a conical flask and place it in a magnetic stirring water bath. Add distilled water to control the substrate concentration at 10% (mass fraction), and stir magnetically to make the raw materials evenly dispersed. Then add cellulase to the conical flask so that the amount of cellulase used in the whole hydrolysis system is 0.07 g / g of the substrate, control the pH to be 5.5, and finally seal the bottle mouth with plastic wrap, and place it under constant temperature conditions and stir continuously for enzymatic hydrolysis to obtain the hydrolysis liquid, that is, the high-efficiency enzymatic hydrolysis of the corncob pretreated by acid and alkali steps to produce glucose is completed;

[0070] Specifically, the conditions for enzymatic hydrolysis treatment under constant temperature conditions are: the hydrolysis temperature is 50 °C, the hydrolysis time is 72 h, and the rotation speed is 500 rpm.

[0071] Specifically, in step (1), the components of the corncob raw material are: the cellulose content is 34% - 38%, the hemicellulose content is 20% - 30%, and the lignin and other contents are 30% - 40%.

[0072] Specifically, in step (1), the screening is carried out with 30 meshes.

[0073] Specifically, in step (7), the pH range is controlled at 5.5, and the acids and alkalis used to adjust the pH are: HCl solution and sodium hydroxide solution.

[0074] Test results of the example: Under the above enzymatic hydrolysis conditions, the concentration of glucose in the hydrolysis liquid is 79.2 g / L, and the glucose yield is 0.868 g / g.

[0075] Example 2

[0076] (1) Raw material preparation: Air-dry, crush and screen the corncob to obtain the treated corncob, and put the obtained treated corncob into a sealed bag for subsequent use;

[0077] (2) Two-step pretreatment with acid and alkali: Weigh the pretreated corncobs from step (1), put them into a conical flask and place it in a water bath. Add sulfuric acid, control the solid-liquid ratio of the raw material suspension to be 1:16, control the sulfuric acid concentration of the raw material suspension to be 2.2%, and control the temperature of the water bath to be 90 °C;

[0078] (3) After the reaction reaches 120 min, collect the reacted corncobs and wash them repeatedly several times until neutral. Collect the washing liquid, and dry the reacted corncobs in an oven at 105 °C to obtain acid-treated corncobs, and store them in an environment of 2-5 °C for subsequent use;

[0079] (4) Weigh the acid-treated corncobs from step (3), put them into a conical flask and place it in a water bath. Add sodium hydroxide and calcium hydroxide, control the solid-liquid ratio of the raw material suspension to be 1:16, control the ratio of sodium hydroxide to calcium hydroxide to be between 5:1, control the total addition amount of sodium hydroxide and calcium hydroxide in the raw material suspension to be 0.09 g / g substrate, and control the temperature of the water bath to be 60 °C;

[0080] (5) After the reaction reaches 3 h, collect the reacted corncobs and wash them repeatedly several times until neutral. Collect the washing liquid, and dry the reacted corncobs in an oven at 105 °C to obtain corncobs pretreated by two steps of acid and alkali, and store them in a low-temperature environment of 2-5 °C for subsequent use;

[0081] (6) After the collected acid-treated liquid and alkali-treated liquid are used for subsequent comprehensive utilization, they are neutralized and discharged;

[0082] (7) Enzymatic hydrolysis: Weigh the corncobs pretreated by two steps of acid and alkali as the substrate for enzymatic hydrolysis, put them into a conical flask and place it in a magnetic stirring water bath. Add distilled water to control the substrate concentration at 4% (mass fraction), and stir magnetically to make the raw materials evenly dispersed. Then add cellulase to the conical flask so that the amount of cellulase used in the whole hydrolysis system is 0.05 g / g substrate, control the pH to be 5.5, and finally seal the bottle mouth with plastic wrap and place it under constant temperature conditions and stir continuously for enzymatic hydrolysis to obtain a hydrolysis solution, that is, the high-efficiency enzymatic hydrolysis of corncobs pretreated by two steps of acid and alkali to produce glucose is completed;

[0083] Specifically, the conditions for enzymatic hydrolysis treatment under constant temperature conditions are: hydrolysis temperature is 50 °C, hydrolysis time is 72 h, and rotation speed is 500 rpm.

[0084] Specifically, in step (1), the components of the corncob raw material are cellulose content of 34% - 38%, hemicellulose content of 20% - 30%, and lignin and other content of 30% - 40%.

[0085] Specifically, in step (1), the screening is carried out with 30 meshes.

[0086] Specifically, in step (7), the pH range is controlled at 5.5, and the acids and bases used to adjust the pH are: HCl solution and sodium hydroxide solution.

[0087] Test results of Example 2: Under the above enzyme hydrolysis conditions, the concentration of glucose in the hydrolysis solution is 29.65 g / L, and the glucose yield is 0.807 g / g.

[0088] Example 3

[0089] (1) Raw material preparation: The corncobs are air-dried, crushed, and screened to obtain processed corncobs, and the obtained processed corncobs are put into a sealed bag for subsequent use;

[0090] (2) Two-step acid and base pretreatment: Weigh the processed corncobs from step (1), put them into a conical flask and place it in a water bath. Add sulfuric acid, control the solid-liquid ratio of the raw material suspension to be 1:16, control the sulfuric acid concentration of the raw material suspension to be 2.2%, and control the temperature of the water bath to be 90 °C;

[0091] (3) After the reaction reaches 120 min, collect the reacted corncobs and wash them repeatedly several times until neutral. Collect the washing liquid, and dry the reacted corncobs in an oven at 105 °C to obtain acid-treated corncobs, and store them in an environment of 2-5 °C for subsequent use;

[0092] (4) Weigh the acid-treated corncobs from step (3), put them into a conical flask and place it in a water bath. Add sodium hydroxide and calcium hydroxide, control the solid-liquid ratio of the raw material suspension to be 1:16, control the ratio of sodium hydroxide to calcium hydroxide to be between 5:1, control the total addition amount of sodium hydroxide and calcium hydroxide in the raw material suspension to be 0.09 g / g substrate, and control the temperature of the water bath to be 60 °C;

[0093] (5) After the reaction reaches 3 h, collect the reacted corncobs and wash them repeatedly several times until neutral. Collect the washing liquid, and dry the reacted corncobs in an oven at 105 °C to obtain corncobs pretreated by two steps of acid and base, and store them in a low temperature environment of 2-5 °C for subsequent use;

[0094] (6) After the collected acid treatment liquid and base treatment liquid are comprehensively utilized later, they are neutralized and discharged;

[0095] (7) Enzymatic hydrolysis: Weigh the corncob after two-step pretreatment with acid and alkali as the substrate for enzymatic hydrolysis, put it into a conical flask, and place it in a magnetic stirring water bath. Add distilled water to control the substrate concentration at 4% (mass fraction). Stir magnetically to evenly disperse the raw materials. Then add cellulase to the conical flask so that the dosage of cellulase in the whole hydrolysis system is 0.05 g / g of the substrate. Control the pH to 5.5. Finally, seal the bottle mouth with plastic wrap and place it under constant temperature conditions with continuous stirring for enzymatic hydrolysis to obtain the hydrolysis solution, thus completing the high-efficiency enzymatic hydrolysis of corncob pretreated by two steps of acid and alkali to produce glucose;

[0096] Specifically, the conditions for enzymatic hydrolysis treatment under constant temperature conditions are: the hydrolysis temperature is 50 °C, the hydrolysis time is 48 h, and the rotation speed is 500 rpm.

[0097] Specifically, in step (1), the composition of the corncob raw material is 34% - 38% cellulose content, 20% - 30% hemicellulose content, and 30% - 40% lignin and others content.

[0098] Specifically, in step (1), the screening is carried out with a 30-mesh sieve.

[0099] Specifically, in step (7), the pH range is controlled at 5.5, and the acids and alkalis used are: HCl solution and sodium hydroxide solution.

[0100] Test results of Example 3: Under the above enzymatic hydrolysis conditions, the glucose concentration in the hydrolysis solution is 25.1 g / L, and the glucose yield is 0.682 g / g.

[0101] Example 4

[0102] (1) Raw material preparation: Air-dry, crush, and screen the corncob to obtain the treated corncob, and put the obtained treated corncob into a sealed bag for subsequent use;

[0103] (2) Two-step pretreatment with acid and alkali: Weigh the treated corncob in step (1), put it into a conical flask, and place it in a water bath. Add sulfuric acid, control the solid-liquid ratio of the raw material suspension to be 1:16, control the sulfuric acid concentration of the raw material suspension to be 2.2%, and control the temperature of the water bath to be 90 °C;

[0104] (3) After the reaction reaches 120 min, collect the reacted corncob and wash it repeatedly several times until it is neutral. Collect the washing solution, and dry the reacted corncob in an oven at 105 °C to obtain the acid-treated corncob, and store it in an environment of 2 - 5 °C for subsequent use;

[0105] (4) Weigh the acid-treated corncob from step (3), put it into a conical flask and place it in a water bath. Add sodium hydroxide and calcium hydroxide, control the solid-liquid ratio of the raw material suspension to be 1:16, control the ratio of sodium hydroxide to calcium hydroxide to be between 5:1, control the total addition amount of sodium hydroxide and calcium hydroxide in the raw material suspension to be 0.09 g / g of the substrate, and control the temperature of the water bath to be 60 °C;

[0106] (5) After the reaction reaches 3 h, collect the reacted corncob and wash it repeatedly several times until it is neutral. Collect the washing liquid, and place the reacted corncob in an oven at 105 °C to dry, obtaining the corncob pretreated by acid and alkali steps. Store it in a low-temperature environment of 2-5 °C for subsequent use;

[0107] (6) After the collected acid treatment liquid and alkali treatment liquid are comprehensively utilized subsequently, neutralize and discharge them;

[0108] (7) Enzymatic hydrolysis: Weigh the corncob pretreated by acid and alkali steps as the substrate for enzymatic hydrolysis, put it into a conical flask and place it in a magnetic stirring water bath. Add distilled water to control the substrate concentration at 6% (mass fraction), and stir magnetically to make the raw materials evenly dispersed. Then add cellulase to the conical flask so that the dosage of cellulase in the whole hydrolysis system is 0.05 g / g of the substrate, control the pH to be 6.0, and finally seal the bottle mouth with plastic wrap and place it under constant temperature conditions and stir continuously for enzymatic hydrolysis to obtain the hydrolysis liquid, that is, complete the high-efficiency enzymatic hydrolysis of corncob pretreated by acid and alkali steps to produce glucose;

[0109] Specifically, the conditions for enzymatic hydrolysis treatment under constant temperature conditions are: hydrolysis temperature is 50 °C, hydrolysis time is 72 h, and rotation speed is 500 rpm.

[0110] Specifically, in step (1), the components of the corncob raw material are: cellulose content 34% - 38%, hemicellulose content 20% - 30%, lignin and other content 30% - 40%.

[0111] Specifically, in step (1), the screening is carried out with 30 meshes.

[0112] Specifically, in step (7), the pH range is controlled at 5.5, and the acids and alkalis used are: HCl solution, sodium hydroxide solution.

[0113] Test results of Example 4: Under the above enzymatic hydrolysis conditions, the concentration of glucose in the hydrolysis liquid is 47.5 g / L, and the glucose yield is 0.866 g / g.

[0114] Example 5

[0115] (1) Raw material preparation: Air-dry, crush and screen the corncob to obtain the treated corncob, and put the obtained treated corncob into a sealed bag for subsequent use;

[0116] (2) Two-step pretreatment with acid and alkali: Weigh the pretreated corncobs from step (1), put them into a conical flask and place it in a water bath. Add sulfuric acid, control the solid-liquid ratio of the raw material suspension to be 1:16, control the sulfuric acid concentration of the raw material suspension to be 2.2%, and control the temperature of the water bath to be 90 °C;

[0117] (3) After the reaction reaches 120 min, collect the reacted corncobs and wash them repeatedly several times until neutral. Collect the washing liquid, and place the reacted corncobs in an oven at 105 °C to dry, obtaining the acid-treated corncobs. Store them in an environment of 2 - 5 °C for subsequent use;

[0118] (4) Weigh the acid-treated corncobs from step (3), put them into a conical flask and place it in a water bath. Add sodium hydroxide and calcium hydroxide, control the solid-liquid ratio of the raw material suspension to be 1:16, control the ratio of sodium hydroxide to calcium hydroxide to be between 5:1, control the total addition amount of sodium hydroxide and calcium hydroxide in the raw material suspension to be 0.09 g / g substrate, and control the temperature of the water bath to be 60 °C;

[0119] (5) After the reaction reaches 3 h, collect the reacted corncobs and wash them repeatedly several times until neutral. Collect the washing liquid, and place the reacted corncobs in an oven at 105 °C to dry, obtaining the corncobs after two-step pretreatment with acid and alkali. Store them in a low-temperature environment of 2 - 5 °C for subsequent use;

[0120] (6) After the collected acid-treated liquid and alkali-treated liquid are used for subsequent comprehensive utilization, neutralize and discharge them;

[0121] (7) Enzymatic hydrolysis: Weigh the corncobs after two-step pretreatment with acid and alkali as the substrate for enzymatic hydrolysis, put them into a conical flask and place it in a magnetic stirring water bath. Add distilled water to control the substrate concentration at 8% (mass fraction). Stir magnetically to make the raw materials evenly dispersed. Then add cellulase to the conical flask so that the amount of cellulase used in the whole hydrolysis system is 0.05 g / g substrate. Control the pH to be 5.5. Finally, seal the bottle mouth with plastic wrap and place it under constant temperature conditions with continuous stirring for enzymatic hydrolysis to obtain the hydrolysis liquid, that is, the high-efficiency enzymatic hydrolysis of corncobs after two-step pretreatment with acid and alkali to produce glucose is completed;

[0122] Specifically, the conditions for enzymatic hydrolysis treatment under constant temperature conditions are: hydrolysis temperature is 50 °C, hydrolysis time is 72 h, and rotation speed is 500 rpm.

[0123] Specifically, in step (1), the components of the corncob raw material are cellulose content of 34% - 38%, hemicellulose content of 20% - 30%, and lignin and other contents of 30% - 40%.

[0124] Specifically, in step (1), the screening is carried out with 30 - mesh.

[0125] Specifically, in step (7), the pH range is controlled at 5.5, and the acids and bases used are: HCl solution and sodium hydroxide solution.

[0126] Test results of Example 5: Under the above enzymatic hydrolysis conditions, the concentration of glucose in the hydrolysis solution is 60.2 g / L, and the glucose yield is 0.824 g / g.

[0127] Example 6

[0128] (1) Raw material preparation: The corncobs are air-dried, crushed, and screened to obtain treated corncobs, which are then put into a sealed bag for subsequent use.

[0129] (2) Two-step acid and base pretreatment: Weigh the treated corncobs from step (1), put them into a conical flask and place it in a water bath. Add sulfuric acid, control the solid-liquid ratio of the raw material suspension to be 1:16, control the sulfuric acid concentration of the raw material suspension to be 2.2%, and control the temperature of the water bath to be 90°C.

[0130] (3) After the reaction reaches 120 min, collect the reacted corncobs and wash them repeatedly several times until neutral. Collect the washing liquid, and dry the reacted corncobs in an oven at 105°C to obtain acid-treated corncobs, which are stored in an environment of 2 - 5°C for subsequent use.

[0131] (4) Weigh the acid-treated corncobs from step (3), put them into a conical flask and place it in a water bath. Add sodium hydroxide and calcium hydroxide, control the solid-liquid ratio of the raw material suspension to be 1:16, control the ratio of sodium hydroxide to calcium hydroxide to be between 5:1, control the total addition amount of sodium hydroxide and calcium hydroxide in the raw material suspension to be 0.09 g / g of the substrate, and control the temperature of the water bath to be 60°C.

[0132] (5) After the reaction reaches 3 h, collect the reacted corncobs and wash them repeatedly several times until neutral. Collect the washing liquid, and dry the reacted corncobs in an oven at 105°C to obtain corncobs pretreated by two-step acid and base, which are stored in a low-temperature environment of 2 - 5°C for subsequent use.

[0133] (6) After the collected acid treatment liquid and base treatment liquid are comprehensively utilized later, they are neutralized and discharged.

[0134] (7) Enzymatic hydrolysis: Weigh the corncob after two-step pretreatment with acid and alkali as the substrate for enzymatic hydrolysis, put it into a conical flask, and place it in a magnetic stirring water bath. Add distilled water to control the substrate concentration at 10% (mass fraction). Stir magnetically to evenly disperse the raw materials. Then add cellulase to the conical flask so that the dosage of cellulase in the whole hydrolysis system is 0.09 g / g of the substrate. Control the pH to 5.5. Finally, seal the bottle mouth with plastic wrap and place it under constant temperature conditions with continuous stirring for enzymatic hydrolysis to obtain the hydrolysis solution, thus completing the high-efficiency enzymatic hydrolysis of corncob pretreated by two steps of acid and alkali to produce glucose;

[0135] Specifically, the conditions for enzymatic hydrolysis treatment under constant temperature conditions are: the hydrolysis temperature is 50 °C, the hydrolysis time is 48 h, and the rotation speed is 500 rpm.

[0136] Specifically, in step (1), the composition of the corncob raw material is: cellulose content 34% - 38%, hemicellulose content 20% - 30%, lignin and other content 30% - 40%.

[0137] Specifically, in step (1), the screening is carried out with a 30-mesh sieve.

[0138] Specifically, in step (7), the pH range is controlled at 5.5, and the acids and alkalis used are: HCl solution and sodium hydroxide solution.

[0139] Test results of Example 6: Under the above enzymatic hydrolysis conditions, the glucose concentration in the hydrolysis solution is 68.6 g / L, and the glucose yield is 0.751 g / g.

[0140] Advantages: The corncobs of Examples 1 - 6 are treated by the two-step pretreatment method with acid and alkali. The calculated hemicellulose removal rate reaches 87.88%, and the lignin removal rate reaches 83.84%.

[0141] Comparative Example 1

[0142] Modified part: In the enzymatic hydrolysis step (step (7) of Example 1), the dosage of cellulase is reduced from 0.07 g / g of the substrate to 0.03 g / g of the substrate, and other conditions remain unchanged.

[0143] Test results of Comparative Example 1: Reducing the dosage of cellulase will lead to a decrease in enzymatic hydrolysis efficiency, and the glucose concentration and yield may decrease. The measured glucose concentration is 78.2 g / L, and the glucose yield is 0.802 g / g.

[0144] Comparative Example 2

[0145] Modified part: In the enzymatic hydrolysis step (step (7) of Example 2), the substrate concentration is increased from 4% (mass fraction) to 12%, and other conditions remain unchanged.

[0146] Detection results of Comparative Example 2: Excessive substrate concentration will lead to a decrease in the contact efficiency between the enzyme and the substrate, and at the same time increase the viscosity of the reaction system, which is not conducive to the diffusion and action of the enzyme. The measured glucose concentration is 28.25 g / L, and the glucose yield is 0.787 g / g.

[0147] Comparative Example 3

[0148] Modified part: In the two-step acid and alkali pretreatment steps (Steps (2)-(5) of Example 3), the sulfuric acid concentration was reduced from 2.2% to 1.5%, and other conditions remained unchanged.

[0149] Detection results of Comparative Example 3: A decrease in sulfuric acid concentration will lead to a weakened acid treatment effect and a reduced degree of depolymerization of cellulose and hemicellulose, thus affecting the efficiency of subsequent enzymatic hydrolysis. The measured glucose concentration is 23.9 g / L, and the glucose yield is 0.639 g / g.

[0150] Comparative Example 4

[0151] Modified part: In the enzymatic hydrolysis step (Step (7) of Example 4), the hydrolysis temperature was reduced from 50 °C to 40 °C, and other conditions remained unchanged.

[0152] Detection results of Comparative Example 4: A decrease in hydrolysis temperature will lead to a decrease in enzyme activity, a slowdown in the enzymatic hydrolysis rate, and a reduction in glucose production efficiency. The measured glucose concentration is 43.3 g / L, and the glucose yield is 0.788 g / g.

[0153] Comparative Example 5

[0154] Modified part: In the enzymatic hydrolysis step (Step (7) of Example 5), the hydrolysis time was shortened from 72 hours to 24 hours, and other conditions remained unchanged.

[0155] Detection results of Comparative Example 5: Shortening the hydrolysis time will lead to incomplete enzymatic hydrolysis reaction, and cellulose cannot be fully hydrolyzed into glucose. The measured glucose concentration is lower than 60.2 g / L, and the glucose yield is 0.799 g / g.

[0156] Comparative Example 6

[0157] Modified part: In the two-step acid and alkali pretreatment steps (Steps (2)-(5) of Example 6), the ratio of sodium hydroxide to calcium hydroxide was changed from 5:1 to 1:1, and other conditions remained unchanged.

[0158] Detection results of Comparative Example 6: Changing the ratio of sodium hydroxide to calcium hydroxide will lead to a change in the alkali treatment effect, affect the separation effect of cellulose and lignin, and thus affect the enzymatic hydrolysis efficiency. The measured glucose concentration is 66.7 g / L, and the glucose yield is 0.737 g / g.

[0159] Summary: By comparing Examples 1-6 with their corresponding Comparative Examples 1-6 respectively, it can be seen that in Examples 1-6, through stepwise acid-base pretreatment, impurities in corncobs were effectively removed, hemicellulose and lignin were removed step by step, and cellulose was enriched in the materials, improving the enzymatic hydrolysis efficiency and glucose yield of cellulose in corncobs. Compared with the single pretreatment method, the present invention optimized the pretreatment conditions, reduced the treatment cost, thus avoiding the impact of harsh conditions on equipment and the environment. At the same time, the hierarchical utilization of cellulose, hemicellulose and lignin was realized, further enhancing the comprehensive utilization value of resources. At the same time, the present invention uses agricultural waste corncobs as raw materials, which can not only solve the problems of resource waste and environmental pollution caused by burning corncobs as rural fuel, but also provide a new technical approach for the efficient utilization of biomass resources and the development of new energy, meeting the concept of green sustainable development and having economic and social benefits. In contrast, in Comparative Examples 1-6, although the parameters of enzyme dosage, substrate concentration, acid-base concentration, hydrolysis temperature and hydrolysis time were adjusted, these adjustments did not achieve the optimization effect of the present invention. On the contrary, the adjustment of these parameters affected the final glucose concentration and yield, resulting in a decrease in enzymatic hydrolysis efficiency and a reduction in glucose production, thus reducing the glucose concentration and yield, further verifying that the stepwise acid-base pretreatment method of the present invention can greatly improve the enzymatic hydrolysis efficiency of cellulose and the glucose yield.

[0160] Although the embodiments of the present invention have been shown and described, for those of ordinary skill in the art, it can be understood that various changes, modifications, substitutions and variations can be made to these embodiments without departing from the principle and spirit of the present invention. The scope of the present invention is defined by the appended claims and their equivalents.

Claims

1. A high-yield enzymatic hydrolysis and saccharification method for stepwise removal of impurities from corncobs by acid and alkali, characterized in that, It includes the following steps: S1. Raw material preparation: Air-dry, crush, and screen corncobs to obtain processed corncobs, and put the obtained processed corncobs into a sealed bag for subsequent use; S2. Acid pretreatment: Weigh the processed corncobs in step S1 according to the formula ratio, put them into a conical flask and place it in a water bath. Add sulfuric acid to react under the conditions of controlling the solid-liquid ratio of the raw material suspension, the sulfuric acid concentration, and the temperature of the water bath; S3. Post-treatment after acid treatment: After the reaction reaches 30 - 120 min, collect the reacted corncobs, wash the corncobs until neutral, collect the washing liquid, and dry and store the reacted corncobs; S4. Alkali pretreatment: Weigh the corncobs after acid treatment in step S3, put them into a conical flask and place it in a water bath. Add sodium hydroxide and calcium hydroxide to react under the conditions of controlling the solid-liquid ratio of the raw material suspension, the ratio of sodium hydroxide to calcium hydroxide, the total addition amount of sodium hydroxide and calcium hydroxide in the raw material suspension, and the temperature of the water bath; S5. Post-treatment after alkali treatment: After the reaction reaches 1 h - 5 h, collect the reacted corncobs, wash the corncobs until neutral, collect the washing liquid, and dry and store the reacted corncobs; S6. Treatment of treatment liquid: After the collected acid treatment liquid and alkali treatment liquid are comprehensively utilized later, neutralize and discharge; S7. Enzymatic hydrolysis: Weigh the corncobs after two-step pretreatment with acid and alkali as the substrate for enzymatic hydrolysis, put them into a conical flask and place it in a magnetic stirring water bath. Add cellulase for enzymatic hydrolysis under the conditions of controlling the substrate concentration, the dosage of cellulase, the pH range, and the constant temperature stirring conditions to obtain a hydrolysis solution, that is, complete the high-efficiency enzymatic hydrolysis of corncobs pretreated with acid and alkali to produce glucose.

2. The high-yield enzymatic hydrolysis and saccharification method for stepwise acid-base impurity removal of corncobs according to claim 1, characterized in that: In the above step S1, the composition of the corncob raw material is: cellulose content 34% - 38%, hemicellulose content 20% - 30%, lignin and other content 30% - 40%.

3. A high-yield enzymatic hydrolysis and saccharification method for stepwise acid-base impurity removal from corncobs according to claim 1, characterized in that: In the above step S1, the screening is carried out with 30 meshes.

4. A high-yield enzymatic hydrolysis and saccharification method for stepwise removal of impurities from corncobs by acid-base treatment according to claim 1, characterized in that: The acid pretreatment conditions in S2 are: control the solid-liquid ratio of the raw material suspension at 1:16 - 1:10, control the sulfuric acid concentration of the raw material suspension at 0.7% - 2.2%, and control the temperature of the water bath at 60 - 90 °C.

5. A high-yield enzymatic hydrolysis and saccharification method for stepwise acid-base impurity removal from corncobs according to claim 1, characterized in that: The post-treatment process after acid treatment in S3: S3.

1. After the reaction reaches 30 - 120 min, collect the reacted corncobs and wash them repeatedly several times until the corncobs are neutral, and collect the washing liquid; S3.

2. Place the reacted corncobs in an oven at 105 °C to dry to obtain corncobs after acid treatment; S3.

3. Store it in an environment of 2 - 5 °C for subsequent use.

6. A high-yield enzymatic hydrolysis and saccharification method for stepwise acid-base impurity removal from corncobs according to claim 1, characterized in that: The alkali pretreatment conditions in S4: control the solid-liquid ratio of the raw material suspension at 1:16 - 1:10, control the ratio of sodium hydroxide to calcium hydroxide between 3:1 - 6:1, control the total addition amount of sodium hydroxide and calcium hydroxide in the raw material suspension at 0.03 - 0.15 g / g substrate, and control the temperature of the water bath at 30 - 60 °C.

7. A high-yield enzymatic hydrolysis and saccharification method for stepwise acid-base impurity removal from corncobs according to claim 1, characterized in that: The post-treatment process after alkali treatment in S5: S5.

1. After the reaction reaches 1 h - 5 h, collect the reacted corncobs and wash them repeatedly several times until neutral, and collect the washing liquid; S5.

2. Place the reacted corncobs in an oven at 105 °C for drying to obtain corncobs pretreated by two steps of acid and alkali. S5.

3. Store it in a low-temperature environment of 2-5 °C for subsequent use.

8. A high-yield enzymatic hydrolysis and saccharification method for stepwise acid-base impurity removal from corncobs according to claim 1, characterized in that: The specific process of enzymatic hydrolysis in S7 is as follows: S7.

1. Weigh the corncobs pretreated by two steps of acid and alkali as the enzymatic hydrolysis substrate, put them into a conical flask and place it in a magnetic stirring water bath. S7.

2. Add distilled water to control the substrate concentration at 4% - 12%, and stir magnetically to evenly disperse the raw materials. S7.

3. Then add cellulase to the conical flask so that the dosage of cellulase in the whole hydrolysis system is 0.01 - 0.05 g / g of substrate, and control the pH range at 4.0 - 6.

0. S7.

4. Finally, seal the bottle mouth with plastic wrap, place it under constant temperature conditions and stir continuously for enzymatic hydrolysis to obtain the hydrolysis solution, that is, complete the efficient enzymatic hydrolysis of corncobs pretreated by two steps of acid and alkali to produce glucose.

9. A high-yield enzymatic hydrolysis and saccharification method for stepwise acid-base impurity removal from corncobs according to claim 8, characterized in that: The conditions for enzymatic hydrolysis treatment in S7 are: the hydrolysis temperature is 40 - 60 °C, the hydrolysis time is 24 - 72 h, and the rotation speed is 400 - 500 rpm.

10. A high-yield enzymatic hydrolysis and saccharification method for stepwise acid-base impurity removal from corncobs according to claim 1, characterized in that: In step S7, the pH range is controlled between 4.0 and 6.0, and the acids and alkalis used to control the pH are hydrochloric acid solution and sodium hydroxide solution.

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