A method of solid state fermentation of cellulase

Abstract

The application discloses a novel solid-state fermentation cellulase method, and belongs to the field of fermentation engineering and enzyme engineering. The application comprises the following steps: selecting an inert carrier modified by acid and alkali liquid, loading the inert carrier in a solid-state fermentation tank, then adding a fermentation medium and a seed culture solution to perform cellulase fermentation. Sterile air is periodically introduced during the fermentation process to strengthen the fermentation process. After the fermentation is completed, a crude enzyme solution is obtained through plate-frame extrusion, and then fresh medium is added to continuously perform multiple batches of cellulase fermentation. The application adopts a novel solid-state fermentation cellulase method, has the advantages of high yield, good stability and simple extraction and separation, and achieves the purpose of continuously performing multiple batches of cellulase fermentation.

Description

Technical Field

[0001] This invention belongs to the fields of fermentation engineering and enzyme engineering, and specifically relates to a novel method for solid-state fermentation of cellulase. Background Technology

[0002] Lignocellulose, as one of the most abundant renewable resources in nature, offers a crucial pathway to addressing the energy crisis and environmental issues by fully utilizing its inexpensive and plentiful biomass resources. Cellulases are a class of enzymes that can degrade lignocellulose into glucose, primarily including endoglucanase (EG), exoglucanase (CBH), and β-glucosidase (CB). However, the high cost and low yield of cellulases limit the development of biomass energy, thus necessitating the development of a highly efficient and economical cellulase production process.

[0003] Currently, research on cellulase fermentation strains mainly focuses on filamentous fungi, such as Penicillium, Trichoderma, and others. Among them, Trichoderma reesei is a filamentous fungus that can produce various cellulases. Due to its high yield, good stability, high enzyme activity, and ease of isolation and purification, it is widely used in industry for cellulase production. Cellulase is an inducible enzyme; lactose, cellobiose, microcrystalline cellulose, etc., can induce microbial strains to produce cellulase. Therefore, selecting a suitable carbon source plays an important role in the production of induced cellulase.

[0004] Cellulase production mainly involves liquid fermentation and solid-state fermentation. Solid-state fermentation is a three-phase bioreactor system with the gas phase as the continuous phase and the liquid phase and its solid substrate as the stationary phase. As a commonly used fermentation method, it has advantages such as high yield, low energy consumption, and low wastewater production. It provides a natural and suitable growth environment for fermenting microorganisms and produces a richer enzyme system. However, using traditional nutrient substrate carriers to adsorb cellulase during solid-state fermentation has problems such as substrate clumping, difficulty in product separation and extraction, and unstable fermentation.

[0005] In the production of enzyme preparations, existing technologies mostly employ liquid fermentation with immobilized cells. For example, the technical solution disclosed in Chinese Invention Patent Application Publication No. [CN109554302] proposes using strains immobilized with sodium alginate for liquid fermentation of feed enzyme preparations, while simultaneously recovering the immobilized cells to restore the culture for enzyme preparation production. However, in actual operation, as fermentation progresses, the fermentation broth becomes very viscous, and the substrate degrades and clumps, resulting in very high energy consumption for stirring in the fermenter. Furthermore, the recovery of immobilized cells after fermentation is cumbersome and prone to contamination.

[0006] To address the above issues, the inert carrier adsorption solid-state fermentation method employed in this invention is a novel fermentation approach with the following advantages: ① It provides a favorable adsorption and growth environment for microorganisms, enhancing mass transfer during solid-state fermentation; ② It enables product separation and extraction, reducing the difficulty of separating cellulase during solid-state fermentation; ③ The fermentation strains can be recycled, and continuous multi-batch fermentation can be used to improve cellulase productivity. Summary of the Invention

[0007] The purpose of this invention is to address the shortcomings of existing technologies by providing a novel method for solid-state fermentation of cellulase, which enables continuous multi-batch fermentation of cellulase.

[0008] The technical solution of the present invention is as follows:

[0009] (1) Modification treatment of the carrier: The inert carrier is cut into cubes with a side length of 5mm-10mm, then soaked in an acid solution, treated in a microwave oven for 5-15min, washed with deionized water until neutral, then soaked in an alkaline solution, treated in a microwave oven for 5-15min, finally washed with deionized water until neutral, and dried at 80℃.

[0010] (2) Preparation of fermentation medium: carbon source inducer 30 g / L, yeast powder 0.25 g / L, peptone 0.75 g / L, urea 0.3 g / L, (NH4)2SO4 1.4 g / L, KH2PO4 2 g / L, CaCl2·2H2O 0.4 g / L, MgSO4·7H2O 0.3 g / L, FeSO4·7H2O 0.005 g / L, MnSO4·4H2O 0.0016 g / L, ZnSO4·7H2O 0.0014 g / L, CoCl2·6H2O 0.002 g / L, Tween 80 1 mL / L.

[0011] (3) Carrier and culture medium filling: Inert carrier is filled into solid fermentation tank at 10-60% of the filling amount, and fermentation culture medium is inoculated into solid fermentation tank. The mass ratio of inert carrier to fermentation culture medium is 1:30-70.

[0012] (4) Fermentation and enzyme production culture: Inoculate the cellulase-producing strain seed liquid at 5-20% into the sterilized fermentation medium, open the air pulsation valve, and set the pulsation pressure intensity to 0.05-0.15 MPa and the pulsation frequency to 1 / 30 min. -1 ~1 / 100min -1 Fermentation culture at 28℃.

[0013] (5) Enzyme solution separation and extraction: After the batch fermentation is completed, the discharge port is opened and the crude enzyme solution is separated by plate and frame squeezing. The inert carrier and strain are retained in the solid fermentation tank for the next batch fermentation.

[0014] (6) Continuous multi-batch fermentation: Add fresh fermentation medium to the solid fermentation tank and ferment again to produce enzymes. After 4 to 7 batches of continuous fermentation, the fermentation is terminated.

[0015] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0016] (1) In this invention, porous media materials are selected as inert carriers and acid-base modified by microwave assistance, which increases the specific surface area of ​​the pores, provides a good attachment and growth environment for microorganisms, and improves the stability of the cells during fermentation.

[0017] (2) The present invention uses an inert carrier to adsorb cellulase for solid fermentation. Compared with liquid fermentation, the gas mass transfer effect is better, the fermentation enzyme activity is higher, and the types of enzyme proteins are more abundant.

[0018] (3) The solid fermentation tank used in this invention is equipped with a pulsation device, which can periodically introduce sterile air during the fermentation process, thereby enhancing the mass transfer process of solid fermentation.

[0019] (4) The inert carrier adsorption solid fermentation cellulase method used in this invention is very simple to separate the cellulase product, without the need for complicated processes, and can minimize product loss.

[0020] (5) The present invention uses multiple batches of cellulase fermentation. After each batch of fermentation is completed, the inert carrier and strain are retained, and the enzyme is produced again by adding fresh culture medium, which greatly improves the production efficiency of cellulase. Attached Figure Description

[0021] Figure 1 This is a schematic diagram of a novel solid-state fermentation cellulase process according to the present invention. Detailed Implementation

[0022] The technical solution of the present invention will be further illustrated below through embodiments.

[0023] Comparative Example: Liquid Fermentation Cellulase

[0024] (1) Seed culture

[0025] After culturing Trichoderma reesei on PDA plates for 6 days, spores on the plates were washed off with 0.1 (v / v) Tween 80 solution to prepare a spore suspension. 1 mL of the spore suspension (10... 8 (Spores / mL) were inoculated into 100mL of seed culture medium and cultured at 28℃, pH 5.0, 180rpm for 72 days.

[0026] The seed culture medium consisted of: glucose 10 g / L, yeast extract 0.25 g / L, peptone 0.75 g / L, urea 0.3 g / L, (NH4)2SO4 1.4 g / L, KH2PO4 2 g / L, CaCl2·2H2O 0.4 g / L, MgSO4·7H2O 0.3 g / L, FeSO4·7H2O 0.005 g / L, MnSO4·4H2O 0.0016 g / L, ZnSO4·7H2O 0.0014 g / L, CoCl2·6H2O 0.002 g / L, and Tween 80 1 mL / L.

[0027] (2) Fermentation culture

[0028] The seed culture was inoculated into 100 mL of fermentation medium at a rate of 10% (v / v), and the culture temperature was 28℃, pH was 5.0, the shaking speed was 180 rpm, and the culture time was 168 h.

[0029] The fermentation medium consisted of: microcrystalline cellulose 30 g / L, yeast extract 0.25 g / L, peptone 0.75 g / L, urea 0.3 g / L, (NH4)2SO4 1.4 g / L, KH2PO4 2 g / L, CaCl2·2H2O 0.4 g / L, MgSO4·7H2O 0.3 g / L, FeSO4·7H2O 0.005 g / L, MnSO4·4H2O 0.0016 g / L, ZnSO4·7H2O 0.0014 g / L, CoCl2·6H2O 0.002 g / L, and Tween 80 1 mL / L.

[0030] (3) Cellulase activity assay

[0031] After fermentation, the fermentation broth was centrifuged at 8000 rpm for 10 min, and the supernatant was collected for measurement. The measured cellulase activity was used as a control, and the cellulase activity was 0.262 FPU / mL.

[0032] Example 1

[0033] (1) Carrier modification treatment

[0034] Polyvinyl alcohol sponge (PPF) was cut into cubes with a side length of 5 mm, then soaked in 1 M hydrochloric acid solution and treated in a microwave oven for 5 min. After being thoroughly washed with deionized water until neutral, it was soaked in 1 M sodium hydroxide solution and treated in a microwave oven for 5 min. Finally, it was washed with deionized water until neutral and dried at 80 °C for 24 h to obtain a microwave-assisted modified inert carrier.

[0035] (2) Seed culture

[0036] The strain and seed culture medium are the same as in the comparative example.

[0037] (3) Fermentation culture

[0038] Using treated polyvinyl alcohol sponge (PPF) as the adsorbent, an inert carrier was loaded into a solid-state fermenter at a loading of 20%, followed by the addition of fermentation medium at a mass ratio of 1:30. Seed culture was then inoculated into the fermentation medium at a rate of 10% (v / v). The fermentation temperature was 28℃, pH 5.0, and the incubation time was 168 h. During fermentation, sterile air was periodically purged at a pulsating pressure of 0.05 MPa and a pulsating frequency of 1 / 60 min. -1 .

[0039] The fermentation medium consisted of: lactose 30 g / L, yeast extract 0.25 g / L, peptone 0.75 g / L, urea 0.3 g / L, (NH4)2SO4 1.4 g / L, KH2PO4 2 g / L, CaCl2·2H2O 0.4 g / L, MgSO4·7H2O 0.3 g / L, FeSO4·7H2O 0.005 g / L, MnSO4·4H2O 0.0016 g / L, ZnSO4·7H2O 0.0014 g / L, CoCl2·6H2O 0.002 g / L, and Tween 80 1 mL / L.

[0040] (4) Fermentation enzyme activity assay

[0041] After fermentation, the fermentation broth was collected by plate and frame compression, centrifuged at 8000 rpm for 10 min, and the supernatant was taken for enzyme activity determination. The PPF adsorption solid fermentation cellulase activity was 0.276 FPU / mL, which was 5.5% higher than that of the control example.

[0042] Example 2

[0043] (1) Carrier modification treatment

[0044] Polyurethane foam (PUF) was cut into cubes with a side length of 10 mm, then immersed in 1.5 M hydrochloric acid solution and treated in a microwave oven for 10 min. After being thoroughly washed with deionized water until neutral, it was immersed in 1.5 M sodium hydroxide solution and treated in a microwave oven for 10 min. Finally, it was washed with deionized water until neutral and dried at 80 °C for 24 h to obtain a microwave-assisted modified inert carrier.

[0045] (2) Seed culture

[0046] The strain and seed culture medium are the same as in the comparative example.

[0047] (3) Fermentation culture

[0048] Treated polyurethane foam (PUF) was used as an inert adsorbent carrier and filled into a solid-state fermenter at a loading of 30%. Fermentation medium was added, with a mass ratio of inert carrier to fermentation medium of 1:60. Seed culture was then inoculated into the fermentation medium at a rate of 5% (v / v). The fermentation temperature was 28℃, the pH was 5.0, and the fermentation time was 168 h. During fermentation, sterile air was periodically purged at a pulsating pressure of 0.15 MPa and a pulsating frequency of 1 / 30 min. -1 .

[0049] The fermentation medium consisted of: microcrystalline cellulose 30 g / L, yeast extract 0.25 g / L, peptone 0.75 g / L, urea 0.3 g / L, (NH4)2SO4 1.4 g / L, KH2PO4 2 g / L, CaCl2·2H2O 0.4 g / L, MgSO4·7H2O 0.3 g / L, FeSO4·7H2O 0.005 g / L, MnSO4·4H2O 0.0016 g / L, ZnSO4·7H2O 0.0014 g / L, CoCl2·6H2O 0.002 g / L, and Tween 80 1 mL / L.

[0050] (4) Fermentation enzyme activity assay

[0051] After fermentation, the fermentation broth was collected by plate and frame compression, centrifuged at 8000 rpm for 10 min, and the supernatant was taken for enzyme activity determination. The cellulase activity of PUF adsorption solid fermentation was 0.310 FPU / mL, which was 18.3% higher than that of the control example.

[0052] Example 3

[0053] (1) Carrier modification treatment

[0054] Carbon black polyurethane foam (PAF) with a pore size of 0.6 mm was cut into cubes with a side length of 10 mm. The cubes were then soaked in a 2 M phosphoric acid solution and microwaved for 15 min. After being thoroughly washed with deionized water until neutral, the cubes were soaked in a 2 M ammonia solution and microwaved for 15 min. Finally, the cubes were washed with deionized water until neutral and dried at 80 °C for 24 h to obtain a microwave-assisted modified inert carrier.

[0055] (2) Seed culture

[0056] The strain and seed culture medium are the same as in the comparative example.

[0057] (3) Fermentation culture

[0058] Treated carbon black polyurethane foam (PAF) was used as an inert adsorbent carrier. The inert carrier was filled into a solid-state fermenter at a loading of 40%, followed by the addition of fermentation medium at a mass ratio of 1:40. Seed culture was then inoculated into the fermentation medium at a rate of 10% (v / v). The fermentation temperature was 28℃, pH 5.0, and the incubation time was 168 h. During fermentation, sterile air was periodically purged at a pulsating pressure of 0.10 MPa and a pulsating frequency of 1 / 90 min. -1 .

[0059] The fermentation medium consisted of: 30 g / L of steam-exploded straw, 0.25 g / L of yeast powder, 0.75 g / L of peptone, 0.3 g / L of urea, 1.4 g / L of (NH4)2SO4, 2 g / L of KH2PO4, 0.4 g / L of CaCl2·2H2O, 0.3 g / L of MgSO4·7H2O, 0.005 g / L of FeSO4·7H2O, 0.0016 g / L of MnSO4·4H2O, 0.0014 g / L of ZnSO4·7H2O, 0.002 g / L of CoCl2·6H2O, and 1 mL / L of Tween 80.

[0060] (4) Fermentation enzyme activity assay

[0061] After fermentation, the fermentation broth was collected by plate and frame compression, centrifuged at 8000 rpm for 10 min, and the supernatant was taken for enzyme activity determination. The cellulase activity of PAF adsorption solid fermentation was 0.368 FPU / mL, which was 40.5% higher than that of the control example.

[0062] Example 4

[0063] (1) Carrier modification treatment

[0064] Carbon black polyurethane foam (PAF) with a pore size of 0.6 mm was cut into cubes with a side length of 8 mm. The cubes were then soaked in 1 M phosphoric acid solution and microwaved for 15 min. After being thoroughly washed with deionized water until neutral, the cubes were soaked in 1 M ammonia solution and microwaved for 15 min. Finally, the cubes were washed with deionized water until neutral and dried at 80 °C for 24 h to obtain microwave-assisted modified inert carriers.

[0065] (2) Seed culture

[0066] The strain and seed culture medium are the same as in the comparative example.

[0067] (3) Continuous multi-batch fermentation culture

[0068] Treated carbon black polyurethane foam (PAF) was used as an inert adsorbent carrier. The inert carrier was filled into a solid-state fermenter at 60% capacity, followed by fermentation medium at a mass ratio of 1:40. Seed culture was then inoculated into the fermentation medium at 15% (v / v). The fermentation temperature was 28℃, pH 5.0, and the incubation time was 168 h. During fermentation, sterile air was periodically purged at a pressure of 0.05 MPa and a pulsation frequency of 1 / 80 min. -1 After fermentation is complete, the fermentation broth is collected by plate and frame compression, retaining the carrier and cells. Fresh fermentation medium is then added, and the culture conditions are the same as those for the first batch. This process is repeated for six batches.

[0069] The fermentation medium consisted of: 30 g / L of steam-exploded straw, 0.25 g / L of yeast powder, 0.75 g / L of peptone, 0.3 g / L of urea, 1.4 g / L of (NH4)2SO4, 2 g / L of KH2PO4, 0.4 g / L of CaCl2·2H2O, 0.3 g / L of MgSO4·7H2O, 0.005 g / L of FeSO4·7H2O, 0.0016 g / L of MnSO4·4H2O, 0.0014 g / L of ZnSO4·7H2O, 0.002 g / L of CoCl2·6H2O, and 1 mL / L of Tween 80.

[0070] (4) Fermentation enzyme activity assay

[0071] The fermentation broth obtained from each batch was collected and centrifuged at 8000 rpm for 10 min. The supernatant was then used for enzyme activity determination. Compared with the control example of cellulase fermentation, the fermentation time of each batch of cellulase adsorbed on the inert carrier was shortened to 120 h during the continuous fermentation of 6 batches. The enzyme activity of each batch remained relatively stable during the fermentation process, with an average enzyme activity of 0.318 FPU / mL per batch. Compared with the control example, the enzyme activity of fermented cellulase was increased by 21.4%.

[0072] Example 5

[0073] (1) Carrier modification treatment

[0074] Carbon black polyurethane foam (PAF) with a pore size of 0.2 mm was cut into cubes with a side length of 5 mm. The cubes were then immersed in 1 M hydrochloric acid solution and microwaved for 10 min. After being thoroughly washed with deionized water until neutral, the cubes were immersed in 1 M sodium hydroxide solution and microwaved for 10 min. Finally, the cubes were washed with deionized water until neutral and dried at 80 °C for 24 h to obtain microwave-assisted modified inert carriers.

[0075] (2) Seed culture

[0076] The strain and seed culture medium are the same as in the comparative example.

[0077] (3) Continuous multi-batch fermentation culture

[0078] Treated carbon black polyurethane foam (PAF) was used as an inert adsorbent carrier. The inert carrier was filled into a solid-state fermenter at a loading of 40%, followed by the addition of fermentation medium at a mass ratio of 1:60. Seed culture was then inoculated into the fermentation medium at a rate of 10% (v / v). The fermentation temperature was 28℃, pH 5.0, and the incubation time was 168 h. During fermentation, sterile air was periodically purged at a pulsating pressure of 0.15 MPa and a pulsating frequency of 1 / 30 min. -1 After fermentation is complete, the fermentation broth is collected by plate and frame compression, retaining the carrier and cells. Fresh fermentation medium is then added, and the culture conditions are the same as those for the first batch. This process is repeated for seven batches.

[0079] The fermentation medium consisted of: microcrystalline cellulose 30 g / L, yeast extract 0.25 g / L, peptone 0.75 g / L, urea 0.3 g / L, (NH4)2SO4 1.4 g / L, KH2PO4 2 g / L, CaCl2·2H2O 0.4 g / L, MgSO4·7H2O 0.3 g / L, FeSO4·7H2O 0.005 g / L, MnSO4·4H2O 0.0016 g / L, ZnSO4·7H2O 0.0014 g / L, CoCl2·6H2O 0.002 g / L, and Tween 80 1 mL / L.

[0080] (4) Fermentation enzyme activity assay

[0081] The fermentation broth obtained from each batch was collected and centrifuged at 8000 rpm for 10 min. The supernatant was then used for enzyme activity determination. Compared with the control example of cellulase fermentation, the fermentation time of solid-state cellulase adsorbed on an inert carrier was shortened to 120 h during the continuous fermentation of 7 batches. The enzyme activity of each batch remained relatively stable during the fermentation process, with an average enzyme activity of 0.290 FPU / mL per batch. Compared with the control example, the enzyme activity of fermented cellulase was increased by 10.7%.

[0082] Example 6

[0083] (1) Carrier modification treatment

[0084] Carbon black polyurethane foam (PAF) with a pore size of 1.5 mm was cut into cubes with a side length of 15 mm, then soaked in 1 M hydrochloric acid solution and treated in a microwave oven for 5 min. After being thoroughly washed with deionized water until neutral, it was soaked in 1 M sodium hydroxide solution and treated in a microwave oven for 5 min. Finally, it was washed with deionized water until neutral and dried at 80 °C for 24 h to obtain a microwave-assisted modified inert carrier.

[0085] (2) Seed culture

[0086] The strain and seed culture medium are the same as in the comparative example.

[0087] (3) Continuous multi-batch fermentation culture

[0088] Treated carbon black polyurethane foam (PAF) was used as an inert adsorbent carrier. The inert carrier was filled into a solid-state fermenter at a loading of 50%, followed by the addition of fermentation medium at a mass ratio of 1:30. Seed culture was then inoculated into the fermentation medium at a rate of 10% (v / v). The fermentation temperature was 28℃, pH 5.0, and the incubation time was 168 h. During fermentation, sterile air was periodically purged at a pulsating pressure of 0.05 MPa and a pulsating frequency of 1 / 100 min. -1 After fermentation is complete, the fermentation broth is collected by plate and frame compression, retaining the carrier and cells. Fresh fermentation medium is then added, and the culture conditions are the same as those for the first batch. This process is repeated for five batches.

[0089] The fermentation medium consisted of: 30 g / L of steam-exploded straw, 0.25 g / L of yeast powder, 0.75 g / L of peptone, 0.3 g / L of urea, 1.4 g / L of (NH4)2SO4, 2 g / L of KH2PO4, 0.4 g / L of CaCl2·2H2O, 0.3 g / L of MgSO4·7H2O, 0.005 g / L of FeSO4·7H2O, 0.0016 g / L of MnSO4·4H2O, 0.0014 g / L of ZnSO4·7H2O, 0.002 g / L of CoCl2·6H2O, and 1 mL / L of Tween 80.

[0090] (4) Fermentation enzyme activity assay

[0091] The fermentation broth obtained from each batch was collected and centrifuged at 8000 rpm for 10 min. The supernatant was then used for enzyme activity determination. Compared with the control example of cellulase fermentation, the fermentation time of solid-state cellulase adsorbed on an inert carrier was shortened to 144 h during the continuous fermentation of 5 batches. The enzyme activity of each batch remained relatively stable during the fermentation process, with an average enzyme activity of 0.305 FPU / mL per batch. Compared with the control example, the enzyme activity of fermented cellulase was increased by 16.4%.

Claims

1. A method for solid-state fermentation of cellulase, comprising the following steps: (1) Cut the inert carrier into cubes with a side length of 5mm-15mm, then soak them in an acid solution, treat them in a microwave oven for 5-15min, wash them with deionized water until neutral, then soak them in an alkaline solution, treat them in a microwave oven for 5-15min, finally wash them with deionized water until neutral, dry them at 80℃ and store them as solid fermentation adsorption carriers for later use. (2) The inert carrier treated in step (1) is filled into a solid fermentation tank in a certain proportion, cellulase fermentation medium is added, sterilized at 121℃ for 20 min, cooled to room temperature, and seed liquid is inoculated into fermentation medium in a certain proportion. (3) During the fermentation process, microorganisms are statically cultured in a solid fermentation tank, and sterile air is periodically introduced during the fermentation period to enhance the fermentation process; (4) After fermentation, open the outlet and filter the crude enzyme solution through plate and frame extrusion. Retain the inert carrier and strain, and then add sterilized fresh fermentation medium to the solid fermentation tank to ferment cellulase in multiple batches.

2. A process for solid state fermentation of cellulases as claimed in claim 1, wherein: The inert carrier mentioned in step (1) includes polyurethane foam (PUF), polyvinyl alcohol foam (PPF), or carbon black polyurethane foam (PAF).

3. A process for solid state fermentation of cellulase according to claim 1, wherein: In step (2), the amount of inert carrier in the solid fermenter is 10-60%, and the mass ratio of inert carrier to fermentation medium is 1:30-70.

4. A process for solid state fermentation of cellulase according to claim 1, wherein: In step (2), the genetically engineered bacteria constructed using filamentous fungi as chassis cells are used as cellulase fermentation strains, and the seed liquid is inoculated into the fermentation medium at a ratio of 5% to 20% v / v.

5. A process for solid state fermentation of cellulase according to claim 1, wherein: In step (2), the fermentation medium uses glucose, lactose, microcrystalline cellulose, sodium carboxymethyl cellulose or steam-exploded straw as an inducer.

6. A process for solid state fermentation of cellulase according to claim 1, characterized in that: In step (3), the solid-state fermenter is equipped with a pressure pulsation device, which periodically introduces sterile air at a pressure of 0.05-0.15 MPa and a pulsation frequency of 1 / 30 min. -1 ~1 / 100min -1 .

7. A process for solid state fermentation of cellulase according to claim 1, wherein: In step (4), a filter screen is installed at the bottom of the solid fermentation tank, and crude cellulase solution is obtained by plate and frame compression filtration.

8. A process for solid state fermentation of cellulase according to claim 1, characterized in that: In step (4), after the first batch of fermentation is completed, the carrier and fermentation strain are retained, and then fresh culture medium is added through a hose connected to the tank for the next batch of fermentation. After 4 to 7 batches of fermentation, the fermentation is terminated.

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