Process for preparing corn bran araboxylan by ammonia fiber expansion and compound enzyme method

Abstract

The invention discloses a process for preparing corn bran arabinoxylan through ammonia fiber expansion and a compound enzyme method, and relates to the technical field of preparation of arabinoxylan, and the corn bran arabinoxylan comprises the following components in parts by weight: 100 parts of dried corn bran, 200-300 parts of ammonia water, 1-3 parts of a compound enzyme preparation and 500-800 parts of a citric acid-sodium citrate buffer solution. According to the method, ammonia fiber expansion pretreatment is adopted to replace traditional steam explosion and strong alkali extraction, so that a lignin-cellulose-hemicellulose cross-linked structure in the corn bran can be deeply destroyed, lignin is fully dissolved, favorable conditions are created for subsequent enzymolysis, meanwhile, high-concentration wastewater is prevented from being generated, the environmental burden is reduced, and the method is suitable for industrial production. Degradation of arabinoxylan is not caused, and the structural integrity of polysaccharide is guaranteed; through the mutual synergistic effect of xylanase, arabinofuranosidase and cellulase in the compound enzyme preparation, compared with traditional double-enzyme combination, arabinose side chains can be hydrolyzed in a targeted mode, and more comprehensive substrate degradation is achieved.

Description

Technical Field

[0001] This invention belongs to the field of arabinoxylan preparation technology, and in particular relates to the process of preparing corn husk arabinoxylan by expanding ammonia cellulose and using a compound enzymatic method. Background Technology

[0002] Arabicaxylan is the most abundant functional polysaccharide in corn husks (accounting for 20%–30% of the dry weight of corn husks). It possesses physiological activities such as lowering blood lipids, regulating intestinal flora, and enhancing immunity, and is widely used in food (e.g., prebiotics, thickeners), pharmaceuticals (e.g., drug carriers), and cosmetics (e.g., moisturizers). With the rapid development of the health industry, the market demand for high-purity, high-yield arabinoxylan continues to grow, and the resource utilization of corn husks as an agricultural byproduct (annual output exceeding 100 million tons) has become a research hotspot in the field of natural product extraction.

[0003] Currently, the extraction processes for arabinoxylan from corn bran mainly include alkaline extraction, enzymatic extraction, and a combination of physicochemical methods. For example, CN117187315A proposes a method for extracting arabinoxylan from corn bran and its application. Using corn bran as raw material, the arabinoxylan product is obtained through steam explosion treatment, alkaline extraction, combined extraction with xylanase and cellulase, ethanol precipitation, drying, and ultra-fine pulverization.

[0004] The above-mentioned patent has the following defects in use:

[0005] During the processing, steam explosion can only destroy the surface structure of corn husks (porosity increase of less than 20%), and cannot penetrate deep into the interior to dissolve lignin, thus limiting the efficiency of subsequent enzymatic hydrolysis. At the same time, the alkaline extraction method (NaOH solution) to extract arabinoxylan by destroying the lignin-cellulose-hemicellulose cross-linking structure of corn husks easily generates a large amount of high-concentration wastewater, polluting the environment and easily degrading arabinoxylan. Although this method uses a composite enzymatic extraction method, it only uses xylanase + cellulase, which cannot hydrolyze the arabinose side chains, resulting in insufficient purity. Therefore, this invention proposes a process for preparing corn husk arabinoxylan by ammonia cellulose expansion and a composite enzymatic method. Summary of the Invention

[0006] This invention provides a process for preparing corn husk arabinoxylan using ammonia cellulose expansion and a composite enzymatic method. By employing ammonia cellulose expansion pretreatment instead of traditional steam explosion and strong alkali extraction, it can deeply disrupt the lignin-cellulose-hemicellulose cross-linked structure within the corn husk, fully dissolving lignin and creating favorable conditions for subsequent enzymatic hydrolysis. Simultaneously, it avoids the generation of high-concentration wastewater, reducing environmental burden, and does not cause degradation of arabinoxylan, ensuring the integrity of the polysaccharide structure. Through the synergistic effect of xylanase, arabinofuranase, and cellulase in the composite enzyme preparation, compared to traditional two-enzyme combinations, it can target... The method involves reactive hydrolysis of the arabinose side chain to achieve more comprehensive substrate degradation. Combined with a two-stage enzyme addition approach, this further enhances enzymatic hydrolysis efficiency, reduces substrate residue, and significantly improves the purity of the arabinoxylan product. A combined separation and purification process using cross-flow membrane filtration and ultrafiltration membrane concentration efficiently removes impurities, small molecule contaminants, and incompletely hydrolyzed substrate from the hydrolysate, while precisely controlling the concentration of the concentrate. Optimized and reasonable subsequent spray drying process parameters ensure thorough product collection and sealed storage, guaranteeing suitable moisture content and high stability for easy subsequent storage and application. In summary, this method solves the problems in the background technology.

[0007] To solve the above-mentioned technical problems, the present invention is achieved through the following technical solution:

[0008] The process for preparing corn husk arabinoxylan by ammonia fiber expansion and compound enzyme method of the present invention includes the following parts by weight: 100 parts by weight of dried corn husk, 200-300 parts by weight of ammonia water, 1-3 parts by weight of compound enzyme preparation, and 500-800 parts by weight of citrate-sodium citrate buffer solution, and includes the following preparation steps:

[0009] S1. Pretreatment of ammonia fiber expansion: Mix dried corn husks with ammonia water and react at 100-120℃ and 0.15-0.2MPa for 30-60 minutes. After cooling to room temperature, filter and wash the filter residue with deionized water until neutral.

[0010] S2, Compound Enzymatic Hydrolysis: Add the filter residue obtained in S1 to the citrate-sodium citrate buffer solution, stir evenly, add the compound enzyme preparation, and enzymatically hydrolyze for 2-4 hours at 50-55℃ and 150-200rpm.

[0011] S3. Separation and purification: Centrifuge the enzyme hydrolysate at 4000-5000 rpm for 10-15 minutes, take the supernatant and filter it through a 0.22 μm cross-flow membrane, then concentrate it to a concentration of 10%-15% using an ultrafiltration membrane with a molecular weight cutoff of 10000 Da;

[0012] S4. Spray drying: The concentrate obtained in S3 is spray dried under the conditions of inlet air temperature of 180-200℃, outlet air temperature of 80-90℃, feed rate of 10-15mL / min and atomization pressure of 0.3MPa to obtain arabinoxylan product.

[0013] S5. Product purity test: Take the arabinoxylan product obtained in S4, weigh a quantitative sample and dissolve it in deionized water to prepare a test solution. Then, use high performance liquid chromatography to determine the purity of arabinoxylan in the test solution.

[0014] Furthermore, the moisture content of the dried corn husk 100 in S1 is 5% to 10%, and the mass concentration of the ammonia water is 26-28%.

[0015] Further, the mixing reaction process in S1 is as follows: dry corn husks and ammonia water are added to a high-pressure reactor at a weight ratio of 1:2.5, stirred evenly, and then the reactor is sealed. The temperature is raised to 105-115℃ and the pressure is maintained at 0.16-0.20MPa for 40-50 minutes.

[0016] Further, the filter residue washing process in S1 is as follows: transfer the filtered filter residue to a washing tank, add deionized water until the filter residue is completely submerged, stir for 5-10 minutes and let stand; pour off the supernatant, repeat the washing 2-3 times, take a filter residue sample every 5 minutes, and use a moisture analyzer to detect the water content until the water content reaches 60-70%.

[0017] Furthermore, the compound enzyme preparation comprises xylanase, arabinofuranase, and cellulase, and the weight ratio of xylanase:arabinofuranase:cellulase is 5:3:2. The xylanase activity is 6000-8000 U / g, the arabinofuranase activity is 3000-4000 U / g, the cellulase activity is 1000-2000 U / g, and the pH of the citrate-sodium citrate buffer solution is 5.0-5.5.

[0018] Furthermore, the enzymatic hydrolysis process in S2 is as follows: first, slowly add the filter residue from S1 into a citrate-sodium citrate buffer solution, and stir at a speed of 150-200 rpm; after the filter residue is completely dispersed, add the compound enzyme preparation in proportion, maintain the temperature at 50-55℃, and take 1 mL of the enzymatic hydrolysate every 30 minutes to detect the reducing sugar content using the DNS method.

[0019] Furthermore, the addition process of the compound enzyme preparation in S2 is as follows: the amount of compound enzyme preparation added is 1-3% of the dry weight of corn husk, and it is added to the enzymatic hydrolysis system in two parts; the first addition is 60% of the total amount, and after stirring for 10 minutes, the remaining 40% is added in the second part, and the stirring speed is maintained at 150-200 rpm.

[0020] Further, the separation and purification process in S3 is as follows: the enzymatic hydrolysate is transferred to a centrifuge tube and centrifuged at 4000-5000 rpm for 10-15 minutes. The supernatant is then filtered through a 0.22 μm cross-flow membrane at an operating pressure of 0.15-0.25 MPa. The filtrate is then concentrated through an ultrafiltration membrane with a molecular weight cutoff of 10000 Da at an operating pressure of 0.25-0.35 MPa until the concentration of the concentrate reaches 10-15%.

[0021] Furthermore, the spray drying process in S4 is as follows: the concentrated liquid is transported to the spray dryer through a peristaltic pump, the inlet air temperature is set to 180-200℃, the outlet air temperature is set to 80-90℃, the feed rate is controlled at 10-15mL / min, and the atomization pressure is maintained at 0.25-0.35MPa; the dried product is collected through a cyclone separator and then sealed and stored.

[0022] Further, the product purity detection procedure in S5 is as follows: Take 0.1g of arabinoxylan product and place it in a 100mL volumetric flask, add deionized water to 2 to 3cm below the mark, shake for 10 to 15 minutes to completely dissolve the product; dilute to 100mL with deionized water, shake well, and then take 5mL of the solution to filter through a 0.22μm organic filter membrane and collect the filtrate; use a high-performance liquid chromatograph to detect the filtrate, using an amino column, a mobile phase of acetonitrile and water at a volume ratio of 75:25, a flow rate of 0.8 to 1.2mL / min, a detection wavelength of 280nm, and an injection volume of 10μL.

[0023] The present invention has the following advantages over the prior art:

[0024] (1) Environmentally friendly and efficient pretreatment: This technical solution replaces the traditional steam explosion and strong alkali extraction by using ammonia fiber expansion pretreatment, which can deeply destroy the lignin-cellulose-hemicellulose cross-linked structure inside the corn husk, fully dissolve the lignin, create favorable conditions for subsequent enzymatic hydrolysis, avoid the generation of high-concentration wastewater, reduce the environmental burden, and will not cause the degradation of arabinoxylan, thus ensuring the integrity of the polysaccharide structure.

[0025] (2) Fully hydrolyzed and highly pure: This technical solution utilizes the synergistic effect of xylanase, arabinofuranase and cellulase in the compound enzyme preparation. Compared with the traditional dual enzyme combination, it can specifically hydrolyze the side chain of arabinose, achieving more comprehensive substrate degradation. Combined with the method of adding enzymes in two stages, it further improves the hydrolysis efficiency, reduces substrate residue, and significantly improves the purity of arabinoxylan products.

[0026] (3) Thorough separation and stable quality: This technical solution uses a combination of cross-flow membrane filtration and ultrafiltration membrane concentration to separate and purify the product. It can efficiently remove impurities, small molecule pollutants and undigested substrates from the enzymatic hydrolysate, accurately control the concentration of the concentrate, optimize the subsequent spray drying process parameters, and ensure that the product is collected thoroughly and sealed for preservation. This ensures that the product has a suitable water content and strong stability, making it easy to store and use in the future.

[0027] Of course, any product implementing this invention does not necessarily need to achieve all of the advantages described above at the same time. Attached Figure Description

[0028] To more clearly illustrate the technical solutions of the embodiments of the present invention, the accompanying drawings used in the description of the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0029] Figure 1 This is a schematic diagram of the preparation process of corn husk arabinoxylan by ammonia fiber expansion and compound enzymatic method according to the present invention. Detailed Implementation

[0030] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention. Specific Implementation Example 1:

[0032] Please see Figure 1 As shown, the process for preparing corn husk arabinoxylan by ammonia fiber expansion and compound enzyme method of the present invention includes the following parts by weight: 100 parts by weight of dried corn husk, 200-300 parts by weight of ammonia water, 1-3 parts by weight of compound enzyme preparation, and 500-800 parts by weight of citric acid-sodium citrate buffer solution, and includes the following preparation steps:

[0033] S1. Pretreatment of ammonia fiber expansion: Mix dried corn husks with ammonia water and react at 100-120℃ and 0.15-0.2MPa for 30-60 minutes. After cooling to room temperature, filter and wash the filter residue with deionized water until neutral.

[0034] S2, Compound Enzymatic Hydrolysis: Add the filter residue obtained in S1 to the citrate-sodium citrate buffer solution, stir evenly, add the compound enzyme preparation, and enzymatically hydrolyze for 2-4 hours at 50-55℃ and 150-200rpm.

[0035] S3. Separation and purification: Centrifuge the enzyme hydrolysate at 4000-5000 rpm for 10-15 minutes, take the supernatant and filter it through a 0.22 μm cross-flow membrane, then concentrate it to a concentration of 10%-15% using an ultrafiltration membrane with a molecular weight cutoff of 10000 Da;

[0036] S4. Spray drying: The concentrate obtained in S3 is spray dried under the conditions of inlet air temperature of 180-200℃, outlet air temperature of 80-90℃, feed rate of 10-15mL / min and atomization pressure of 0.3MPa to obtain arabinoxylan product.

[0037] S5. Product purity test: Take the arabinoxylan product obtained in S4, weigh a quantitative sample and dissolve it in deionized water to prepare a test solution. Then, use high performance liquid chromatography to determine the purity of arabinoxylan in the test solution.

[0038] Among them, the moisture content of dried corn husks 100 in S1 is 5% to 10%, and the mass concentration of ammonia water is 26-28%.

[0039] The mixing reaction process in S1 is as follows: dry corn husks and ammonia water are added to a high-pressure reactor at a weight ratio of 1:2.5, stirred evenly, and then the reactor is sealed. The temperature is raised to 105-115℃ and the pressure is maintained at 0.16-0.20MPa for 40-50 minutes.

[0040] During the reaction, the temperature is raised to 105-115℃, which accelerates the molecular motion rate and increases the penetration efficiency of ammonia into the micropores of corn husks. The pressure is 0.16-0.20 MPa. The high-pressure environment makes it easier for ammonia to break through the lignin-cellulose-hemicellulose cross-linking network of corn husks. Through swelling, it destroys the cellulose crystal zone, degrades some lignin, and releases the encapsulation and binding of arabinoxylan. This enhances the destructive effect of ammonia on the dense structure of corn husks and exposes the arabinoxylan component.

[0041] The filter residue washing process in S1 is as follows: transfer the filtered filter residue to a washing tank, add deionized water until the filter residue is completely submerged, stir for 5-10 minutes and let stand; pour off the supernatant, repeat the washing 2-3 times, take a filter residue sample every 5 minutes, and use a moisture analyzer to test the water content until the water content reaches 60-70%.

[0042] During the filter residue washing process, taking a sample of the filter residue every 5 minutes allows for precise control of the washing endpoint, ensuring a suitable moisture content in the filter residue. This provides an optimized substrate state for subsequent enzymatic hydrolysis (excessive moisture content dilutes the enzymatic hydrolysis system, reducing reaction efficiency; excessive moisture content results in poor substrate dispersibility, affecting enzyme-substrate contact. Maintaining a moisture content in the 60-70% range balances substrate dispersibility and enzyme concentration, improving the enzymatic hydrolysis effect). In this embodiment, the moisture analyzer is designed based on the loss on drying method, and the core process is as follows:

[0043] Initial weighing: Take a quantitative sample of filter residue and record the initial weight;

[0044] Heating and drying: The moisture in the sample is rapidly evaporated by infrared / halogen heating;

[0045] Weight tracking: Monitor sample weight changes in real time until the weight stabilizes (moisture has completely evaporated);

[0046] Results calculation: The moisture content is calculated by the difference between the initial weight and the dried weight (moisture content = (initial weight - dried weight) / initial weight).

[0047] The compound enzyme preparation includes xylanase, arabinofuranase, and cellulase, with a weight ratio of xylanase:arabinofuranase:cellulase of 5:3:2. The xylanase activity is 6000-8000 U / g, the arabinofuranase activity is 3000-4000 U / g, the cellulase activity is 1000-2000 U / g, and the pH of the citrate-sodium citrate buffer is 5.0-5.5.

[0048] The xylanase in the compound enzyme preparation is derived from Aspergillus niger (EC3.2.1.8), the arabinofuranase is derived from Aspergillus oryzae (EC3.2.1.55), and the cellulase is derived from Trichoderma reesei (EC3.2.1.4).

[0049] Xylanase (6000-8000U / g): High activity is required to rapidly break the arabinoxylan backbone, but excessively high activity will significantly increase the cost of enzyme preparations. This range can achieve a balance between efficiency and cost.

[0050] Arabinofuranosylase (3000-4000U / g): The activity needs to match the side chain treatment requirements of xylanase to ensure that the side chain cleavage rate does not lag behind the main chain degradation and to avoid affecting the overall efficiency due to side chain residue.

[0051] Cellulase (1000-2000 U / g): Only sufficient activity is needed to disrupt the cellulose network to expose the substrate. Excessive activity provides no additional benefit and increases costs. This range is sufficient for barrier removal requirements.

[0052] The three enzymes form a highly efficient synergistic system through functional complementarity:

[0053] The pre-activation function of cellulase: First, it breaks down the cellulose network surrounding arabinoxylan, breaking down the dense fibrous structure and providing a spatial channel for the subsequent enzymatic action;

[0054] Side chain treatment of arabinofuranosaccharidase: Remove the arabinose side chain on the arabinoxylan backbone to eliminate the steric hindrance of the side chain to the backbone, making it easier for xylanase to access the β-1,4 glycosidic bonds of the backbone.

[0055] Xylanase degrades the backbone: breaks the arabinoxylan backbone, releasing target products such as xylooligosaccharides or xylose.

[0056] The three work synergistically to effectively overcome the limitations of single enzymes or dual enzyme combinations in substrate contact and steric hindrance clearance, thereby improving the extraction efficiency and purity of arabinoxylan.

[0057] The pH of the citrate-sodium citrate buffer solution is set to 5.0-5.5, based on the optimal catalytic environment requirements of the enzyme.

[0058] This pH range falls within the common optimal pH range for xylanase, arabinofuranase, and cellulase, ensuring that all three enzymes maintain high catalytic activity and preventing enzyme activity decreases or becomes inactive due to pH deviation, thus ensuring the stable performance of their synergistic effect.

[0059] The enzymatic hydrolysis process in S2 is as follows: First, slowly add the filter residue from S1 into a citrate-sodium citrate buffer solution and stir at 150-200 rpm. After the filter residue is completely dispersed, add the compound enzyme preparation in proportion, maintain the temperature at 50-55℃, and take 1 mL of the enzymatic hydrolysate every 30 minutes to detect the reducing sugar content using the DNS method.

[0060] The DNS method, short for 3,5-dinitrosalicylic acid method, is based on the redox reaction between the reducing properties of reducing sugars (containing free aldehyde or ketone groups) and the DNS reagent.

[0061] Under alkaline conditions, reducing sugars (such as xylose and glucose) reduce DNS reagent (yellow) to brownish-red 3-amino-5-nitrosalicylic acid, while simultaneously being oxidized to sugar-acid compounds. The color intensity of the reaction product is linearly positively correlated with the concentration of reducing sugar. By measuring the absorbance at a specific wavelength (usually 540 nm or 550 nm) using a spectrophotometer and combining this with a pre-plotted reducing sugar standard curve, the content of reducing sugar in the sample can be calculated.

[0062] In this embodiment, considering the scenario of "taking 1 mL of enzymatic hydrolysate every 30 minutes" in the enzymatic hydrolysis process, the specific operating steps of the DNS method are as follows:

[0063] Sample pretreatment: Take 1 mL of enzymatic hydrolysate and dilute it with an appropriate amount of deionized water (adjust the dilution factor according to the expected reducing sugar concentration to avoid the absorbance exceeding the linear range).

[0064] Reagent mixing: Add an equal volume (or a preset ratio) of DNS reagent to the diluted sample and mix thoroughly;

[0065] Heating reaction: Place the mixture in a boiling water bath and heat for 5-10 minutes to allow the redox reaction to proceed fully;

[0066] Cooling and volume adjustment: After removing the reaction solution, quickly cool it to room temperature and add deionized water to adjust the volume to a fixed volume (e.g., 10 mL).

[0067] Colorimetric determination: The absorbance of the reaction solution was measured at a wavelength of 540 nm using a spectrophotometer;

[0068] Concentration calculation: Based on the pre-plotted reducing sugar standard curve (using glucose as the standard, a linear regression equation between absorbance and concentration is established), the reducing sugar content in the enzymatic hydrolysate is calculated by substituting the sample absorbance value.

[0069] The core value of the DNS method in this enzymatic hydrolysis process lies in real-time monitoring of the hydrolysis process and quality control, specifically reflected in:

[0070] Dynamic monitoring of the process: Samples are taken every 30 minutes to test the rate of enzymatic hydrolysis by observing the trend of changes in reducing sugar content (such as rapid increase in the early stage and stabilization in the later stage).

[0071] Determination of reaction endpoint: When the reducing sugar content does not increase significantly in multiple consecutive tests, it indicates that the enzymatic hydrolysis reaction has reached equilibrium, and the reaction can be terminated in time to avoid waste of enzyme preparation or excessive degradation of product;

[0072] Process parameter optimization: By monitoring the amount of reducing sugar generated at different time points, parameters such as enzymatic hydrolysis temperature, pH value, and enzyme dosage can be optimized in reverse to improve the degradation efficiency of arabinoxylan.

[0073] Batch stability control: Ensure the consistency of reducing sugar content in different batches of enzymatic hydrolysis products, providing a stable raw material basis for subsequent separation and purification steps.

[0074] The addition process of the compound enzyme preparation in S2 is as follows: the amount of compound enzyme preparation added is 1-3% of the dry weight of corn husk, and it is added to the enzymatic hydrolysis system in two parts; the first addition is 60% of the total amount, and after stirring for 10 minutes, the remaining 40% is added in the second part, and the stirring speed is maintained at 150-200 rpm.

[0075] The purpose of adding the compound enzyme preparation in two separate applications:

[0076] Optimize the initial enzyme-substrate contact: avoid adding a large amount at once, which can lead to localized enzyme aggregation (such as adsorption on specific areas of the substrate surface), reduce ineffective enzyme waste, and ensure that the enzyme is evenly distributed in the system;

[0077] Adapting to dynamic substrate changes: During enzymatic hydrolysis, the substrate structure gradually loosens (cellulose network is destroyed, arabinoxylan sites are exposed). Secondary addition can specifically replenish the amount of enzyme. Utilizing the newly exposed reaction sites, the newly added enzyme can precisely target these newly exposed sites, maintaining catalytic activity in the middle and later stages of the reaction, avoiding premature reaction stagnation, thus accelerating the overall reaction rate of the enzymatic hydrolysis process, significantly increasing the production of reducing sugars (such as xylose and xylooligosaccharides), and optimizing the enzymatic hydrolysis efficiency.

[0078] Maintaining system stability: reducing non-specific inactivation caused by excessively high initial enzyme concentration (such as intermolecular interactions of enzymes) and prolonging the effective action time of the enzyme.

[0079] The separation and purification process in S3 is as follows: the enzymatic hydrolysate is transferred to a centrifuge tube and centrifuged at 4000-5000 rpm for 10-15 minutes. The supernatant is then filtered through a 0.22 μm cross-flow membrane at an operating pressure of 0.15-0.25 MPa. The filtrate is then concentrated through an ultrafiltration membrane with a molecular weight cutoff of 10000 Da at an operating pressure of 0.25-0.35 MPa until the concentration of the concentrate reaches 10-15%.

[0080] Centrifuge (4000-5000 rpm, 10-15 minutes).

[0081] Principle: By utilizing the difference in centrifugal force, the solid and liquid components in the enzymatic hydrolysate are separated (the solid residue settles at the bottom of the tube due to its high density, while the supernatant contains the soluble components).

[0082] Function: Removes insoluble solid residues (such as undegraded corn husk fragments, cellulose residues, and large particle precipitates generated during enzymatic hydrolysis) from the enzymatic hydrolysate; reduces the load on subsequent membrane filtration, decreases the risk of membrane clogging, and extends membrane lifespan.

[0083] 0.22μm crossflow membrane filtration (operating pressure 0.15-0.25MPa)

[0084] Principle: The cross-flow filtration mode (the feed liquid flows parallel to the membrane surface) is adopted. Through the sieving effect of the membrane with a pore size of 0.22μm, particles larger than the pore size are intercepted, while reducing the deposition and fouling on the membrane surface.

[0085] Function: Further removes tiny solid particles (such as submicron residues that have not been centrifuged) from the supernatant; retains some large colloidal impurities (such as undeactivated enzyme proteins and large polysaccharide colloids); improves the clarity of the filtrate, providing a pure feed liquid for subsequent ultrafiltration concentration.

[0086] 10000Da ultrafiltration membrane concentration (operating pressure 0.25-0.35MPa)

[0087] Principle: Based on the molecular weight cutoff characteristics of the membrane, components with a molecular weight ≥10000 Da are retained (assuming the target product arabinoxylan has a molecular weight ≥10000 Da), while small molecule impurities (such as monosaccharides, inorganic salts, and small molecule byproducts) are allowed to permeate through the membrane, and product concentration is achieved through pressure drive.

[0088] Functions: Product concentration: By retaining the target product and discharging the small molecule filtrate, the concentration of arabinoxylan is increased to 10-15%; Small molecule impurity removal: Monosaccharides (such as xylose and glucose), inorganic salts, and small molecule degradation products generated during enzymatic hydrolysis are removed to further purify the target product; Selective separation: The arabinoxylan within the target molecular weight range is retained, and interference from low molecular weight byproducts is reduced.

[0089] The spray drying process in S4 is as follows: the concentrate is delivered to the spray dryer via a peristaltic pump, the inlet air temperature is set to 180-200℃, the outlet air temperature is set to 80-90℃, the feed rate is controlled at 10-15mL / min, and the atomization pressure is maintained at 0.25-0.35MPa; the dried product is collected by a cyclone separator and then sealed for storage.

[0090] In this embodiment, the S4 spray drying process utilizes five core steps: stable delivery of concentrated liquid via a peristaltic pump → atomization into tiny droplets → rapid hot air drying → powder collection via a cyclone separator → sealed storage. This process transforms arabinoxylan concentrate into a solid powder product.

[0091] Among them, atomization into tiny droplets: using an atomization pressure of 0.25-0.35MPa (controlling droplet size: too high pressure → droplets are too small and easily carried away by hot air, reducing the collection rate; too low pressure → droplets are too large, resulting in insufficient drying and powder agglomeration), the concentrate is broken into uniform droplets with a diameter of 10-100μm through a nozzle (or centrifuge disc), which significantly increases the specific surface area of ​​the droplets and provides a basis for rapid drying;

[0092] Hot air rapid drying: Using a co-flow (or counter-flow) hot air drying mode, the incoming air at 180-200℃ (providing the sensible heat required for drying and accelerating the evaporation of moisture on the droplet surface; this temperature range is below the thermal degradation threshold of arabinoxylan to avoid product structure damage) comes into contact with the atomized droplets. Through mass and heat transfer, the moisture on the droplet surface evaporates instantly (the surface temperature is maintained near the wet-bulb temperature to avoid overheating of the product), and the internal moisture gradually diffuses to the surface to complete the drying process, forming a loose powder;

[0093] The cyclone separator collects the powder: After drying, the powder enters the cyclone separator with hot air (outlet temperature 80-90℃, reflecting the drying endpoint: below 80℃ may cause the product to have excessive moisture (easily absorb moisture and clump); above 90℃ may cause the product to overheat and discolor or degrade the active ingredients). With the combined action of centrifugal force and gravity, the powder, due to its high density, settles to the bottom of the separator and is collected, while the hot air is discharged from the top. Sealed storage maintains product stability by isolating it from air and moisture.

[0094] The purity testing procedure in S5 is as follows: Take 0.1g of arabinoxylan product and place it in a 100mL volumetric flask. Add deionized water to 2 to 3cm below the mark and shake for 10 to 15 minutes to completely dissolve the product. Dilute to 100mL with deionized water, shake well, and then take 5mL of the solution and filter it through a 0.22μm organic filter membrane. Collect the filtrate. Detect the filtrate using a high-performance liquid chromatograph. Use an amino column, a mobile phase of acetonitrile and water at a volume ratio of 75:25, a flow rate of 0.8 to 1.2mL / min, a detection wavelength of 280nm, and an injection volume of 10μL.

[0095] The S5 process achieves quantitative analysis of the purity of arabinoxylan products through four core steps: sample dissolution, volume adjustment, filtration, and high-performance liquid chromatography (HPLC) detection.

[0096] Sample pretreatment principle: Through dissolution, volume adjustment, and filtration, solid products are transformed into homogeneous, impurity-free liquid samples, eliminating physical interference and meeting the injection requirements for HPLC detection;

[0097] HPLC detection principle: The specific retention of carbohydrate compounds by an amino column, combined with the polarity adjustment of the acetonitrile-water mobile phase, achieves chromatographic separation of the target component arabinoxylan from impurities; the signal intensity of the target peak is detected at a wavelength of 280 nm, and the purity is calculated by comparing it with the standard (based on external standard method or area normalization method).

[0098] In the high-performance liquid chromatography (HPLC) detection process, an amino column is selected. The amino functional groups form hydrogen bonds with saccharide compounds (arabinoxylan and its components), achieving effective separation of the target component from impurities. It is suitable for normal-phase / reversed-phase separation of sugars. A high proportion of acetonitrile provides a weakly polar environment, enhancing the retention capacity of the amino column for sugars. The mobile phase is a solution of acetonitrile and water at a volume ratio of 75:25. Adjusting the water ratio optimizes the resolution, ensuring complete separation of the target peak from the impurity peak. The flow rate is set to 0.8 to 1.2 m / s. L / min, balancing separation efficiency and analysis time: too low a flow rate leads to peak diffusion and decreased resolution; too high a flow rate results in insufficient separation and inaccurate integration due to peak overlap; the detection wavelength is set to 280 nm to specifically detect the conjugated structure or side chain groups in arabinoxylan, suppressing interference from non-target impurities (such as small molecules without conjugated structures) and improving detection specificity; the injection volume is 10 μL, controlled within the column capacity range, to avoid peak overload (tailing, flat-topped peaks), ensure the linear relationship between peak area and concentration, and improve quantitative accuracy.

[0099] In the specific implementation process, 100g of dried corn husks (9% moisture content), 250g of 25% ammonia water, 2g of compound enzyme preparation (xylanase: arabinofuranase: cellulase = 5:3:2, total activity 12000U / g) and 600mL of pH 5.2 citrate-sodium citrate buffer solution were selected.

[0100] Steps: S1. Mix dried corn husks with ammonia water and react at 110℃ and 0.18MPa for 45 minutes. After cooling to room temperature, filter and wash the filter residue with deionized water until neutral (pH 7.0).

[0101] S2. Add the filter residue obtained in S1 to the citrate-sodium citrate buffer solution, stir evenly, add the compound enzyme preparation, and enzymatically hydrolyze for 3 hours at 52℃ and 180rpm.

[0102] S3. Centrifuge the enzyme hydrolysate at 4500 rpm for 12 minutes, take the supernatant and filter it through a 0.22 μm cross-flow membrane (0.2 MPa), then concentrate it to a concentration of 12% using an ultrafiltration membrane with a molecular weight cutoff of 10000 Da.

[0103] S4. The concentrated liquid obtained in S3 is spray-dried under the conditions of inlet air temperature of 190℃, outlet air temperature of 85℃, feed rate of 12mL / min and atomization pressure of 0.3MPa to obtain arabinoxylan product.

[0104] S5. Take the arabinoxylan product obtained in S4, weigh a quantitative sample and dissolve it in deionized water to prepare a test solution. Then, use high performance liquid chromatography to determine the purity of arabinoxylan in the test solution.

[0105] The process parameters and results of Example 1 are shown in Table 1:

[0106] Specific Implementation Example 2:

[0108] In a preferred embodiment, 100g of dried corn husks (9% moisture content), 250g of 28% ammonia water, 1.8g of a compound enzyme preparation (xylanase: arabinofuranase: cellulase = 5:3:2, total activity 12000U / g) and 600mL of pH 5.2 citrate-sodium citrate buffer solution are selected.

[0109] Steps: S1. Mix dried corn husks with ammonia water and react at 110℃ and 0.18MPa for 50 minutes. After cooling to room temperature, filter and wash the filter residue with deionized water until neutral (pH 7.0).

[0110] S2. Add the filter residue obtained in S1 to the citrate-sodium citrate buffer solution, stir evenly, add the compound enzyme preparation, and enzymatically hydrolyze for 3 hours at 52℃ and 180rpm.

[0111] S3. Centrifuge the enzyme hydrolysate at 4500 rpm for 12 minutes, take the supernatant and filter it through a 0.22 μm cross-flow membrane (0.2 MPa), then concentrate it to a concentration of 12% using an ultrafiltration membrane with a molecular weight cutoff of 10000 Da.

[0112] S4. The concentrated liquid obtained in S3 is spray-dried under the conditions of inlet air temperature of 190℃, outlet air temperature of 85℃, feed rate of 12mL / min and atomization pressure of 0.3MPa to obtain arabinoxylan product.

[0113] S5. Take the arabinoxylan product obtained in S4, weigh a quantitative sample and dissolve it in deionized water to prepare a test solution. Then, use high performance liquid chromatography to determine the purity of arabinoxylan in the test solution.

[0114] The process parameters and results of Example 2 are shown in Table 2:

[0115] Specific Implementation Example 3:

[0117] In a preferred embodiment, 100g of dried corn husks (9% moisture content), 250g of 25% ammonia water, 2g of compound enzyme preparation (xylanase: arabinofuranase: cellulase = 5:3:2, total activity 12000U / g) and 600mL of pH 5.0 citrate-sodium citrate buffer solution are selected.

[0118] Steps: S1. Mix dried corn husks with ammonia water and react at 110℃ and 0.18MPa for 45 minutes. After cooling to room temperature, filter and wash the filter residue with deionized water until neutral (pH 7.0).

[0119] S2. Add the filter residue obtained in S1 to the citrate-sodium citrate buffer solution, stir evenly, add the compound enzyme preparation, and enzymatically hydrolyze for 3.5 hours at 52℃ and 180rpm.

[0120] S3. Centrifuge the enzyme hydrolysate at 4500 rpm for 12 minutes, take the supernatant and filter it through a 0.22 μm cross-flow membrane (0.2 MPa), then concentrate it to a concentration of 12% using an ultrafiltration membrane with a molecular weight cutoff of 10000 Da.

[0121] S4. The concentrated liquid obtained in S3 is spray-dried under the conditions of inlet air temperature of 200℃, outlet air temperature of 85℃, feed rate of 12mL / min and atomization pressure of 0.3MPa to obtain arabinoxylan product.

[0122] S5. Take the arabinoxylan product obtained in S4, weigh a quantitative sample and dissolve it in deionized water to prepare a test solution. Then, use high performance liquid chromatography to determine the purity of arabinoxylan in the test solution.

[0123] The process parameters and results of Example 3 are shown in Table 3:

[0124]

[0125] This invention overcomes the shortcomings of existing technologies by combining ammonia fiber expansion pretreatment with synergistic enzymatic hydrolysis. Example results show that this process achieves an arabinoxylan yield of ≥25%, a purity of ≥90%, and reduces wastewater discharge by 80%.

[0126] The preferred embodiments of the present invention disclosed above are merely illustrative of the invention. These preferred embodiments do not exhaustively describe all details, nor do they limit the invention to the specific implementations described. Clearly, many modifications and variations can be made based on the content of this specification. This specification selects and specifically describes these embodiments to better explain the principles and practical applications of the invention, thereby enabling those skilled in the art to better understand and utilize the invention. The invention is limited only by the claims and their full scope and equivalents.

Claims

1. A process for preparing corn husk arabinoxylan by ammonia fiber expansion and compound enzymatic method, characterized in that, The mixture comprises the following components by weight: 100 parts dried corn husks, 200-300 parts ammonia water, 1-3 parts compound enzyme preparation, and 500-800 parts citrate-sodium citrate buffer solution. The preparation steps are as follows: S1. Pretreatment of ammonia fiber expansion: Mix dried corn husks with ammonia water and react at 100-120℃ and 0.15-0.2MPa for 30-60 minutes. After cooling to room temperature, filter and wash the filter residue with deionized water until neutral. S2, Compound Enzymatic Hydrolysis: Add the filter residue obtained in S1 to the citrate-sodium citrate buffer solution, stir evenly, add the compound enzyme preparation, and enzymatically hydrolyze for 2-4 hours at 50-55℃ and 150-200rpm. S3. Separation and purification: Centrifuge the enzyme hydrolysate at 4000-5000 rpm for 10-15 minutes, take the supernatant and filter it through a 0.22 μm cross-flow membrane, then concentrate it to a concentration of 10%-15% using an ultrafiltration membrane with a molecular weight cutoff of 10000 Da; S4. Spray drying: The concentrate obtained in S3 is spray dried under the conditions of inlet air temperature of 180-200℃, outlet air temperature of 80-90℃, feed rate of 10-15mL / min and atomization pressure of 0.3MPa to obtain arabinoxylan product. S5. Product purity test: Take the arabinoxylan product obtained in S4, weigh a quantitative sample and dissolve it in deionized water to prepare a test solution. Then, use high performance liquid chromatography to determine the purity of arabinoxylan in the test solution.

2. The process for preparing corn husk arabinoxylan by expanding ammonia fiber and using a composite enzyme method according to claim 1, characterized in that, The dried corn husks 100 in S1 have a moisture content of 5% to 10%, and the ammonia water has a mass concentration of 26-28%.

3. The process for preparing corn husk arabinoxylan by expanding ammonia fiber and using a composite enzyme method according to claim 1, characterized in that, The mixing reaction process in S1 is as follows: dry corn husks and ammonia water are added to a high-pressure reactor at a weight ratio of 1:2.5, stirred evenly, and then the reactor is sealed. The temperature is raised to 105-115℃ and the pressure is maintained at 0.16-0.20MPa for 40-50 minutes.

4. The process for preparing corn husk arabinoxylan by expanding ammonia fiber and using a composite enzyme method according to claim 1, characterized in that, The filter residue washing process in S1 is as follows: transfer the filtered filter residue to a washing tank, add deionized water until the filter residue is completely submerged, stir for 5-10 minutes and let stand; pour off the supernatant, repeat the washing 2-3 times, take a filter residue sample every 5 minutes, and use a moisture analyzer to detect the water content until the water content reaches 60-70%.

5. The process for preparing corn husk arabinoxylan by expanding ammonia fiber and using a composite enzyme method according to claim 1, characterized in that, The compound enzyme preparation comprises xylanase, arabinofuranase, and cellulase, with a weight ratio of xylanase:arabinofuranase:cellulase of 5:3:

2. The xylanase activity is 6000-8000 U / g, the arabinofuranase activity is 3000-4000 U / g, and the cellulase activity is 1000-2000 U / g. The pH of the citrate-sodium citrate buffer solution is 5.0-5.

5.

6. The process for preparing corn husk arabinoxylan by expanding ammonia fiber and using a composite enzyme method according to claim 1, characterized in that, The enzymatic hydrolysis process in S2 is as follows: First, slowly add the filter residue from S1 into a citrate-sodium citrate buffer solution, stirring at 150-200 rpm; after the filter residue is completely dispersed, add the compound enzyme preparation in proportion, maintain the temperature at 50-55℃, and take 1 mL of the enzymatic hydrolysate every 30 minutes to detect the reducing sugar content using the DNS method.

7. The process for preparing corn husk arabinoxylan by expanding ammonia fiber and using a composite enzyme method according to claim 1, characterized in that, The addition process of the compound enzyme preparation in S2 is as follows: the amount of compound enzyme preparation added is 1-3% of the dry weight of corn husk, and it is added to the enzymatic hydrolysis system in two parts; the first addition is 60% of the total amount, and after stirring for 10 minutes, the remaining 40% is added in the second part, and the stirring speed is maintained at 150-200 rpm.

8. The process for preparing corn husk arabinoxylan by expanding ammonia fiber and using a composite enzyme method according to claim 1, characterized in that, The separation and purification process in S3 is as follows: the enzymatic hydrolysate is transferred to a centrifuge tube and centrifuged at 4000-5000 rpm for 10-15 minutes. The supernatant is then filtered through a 0.22 μm cross-flow membrane at an operating pressure of 0.15-0.25 MPa. The filtrate is then concentrated through an ultrafiltration membrane with a molecular weight cutoff of 10000 Da at an operating pressure of 0.25-0.35 MPa until the concentration of the concentrate reaches 10-15%.

9. The process for preparing corn husk arabinoxylan by expanding ammonia fiber and using a composite enzyme method according to claim 1, characterized in that, The spray drying process in S4 is as follows: the concentrated liquid is transported to the spray dryer through a peristaltic pump, the inlet air temperature is set to 180-200℃, the outlet air temperature is set to 80-90℃, the feed rate is controlled at 10-15mL / min, and the atomization pressure is maintained at 0.25-0.35MPa; the dried product is collected through a cyclone separator and then sealed and stored.

10. The process for preparing corn husk arabinoxylan by expanding ammonia fiber and using a composite enzyme method according to claim 1, characterized in that, The product purity detection procedure in S5 is as follows: Take 0.1g of arabinoxylan product and place it in a 100mL volumetric flask. Add deionized water to 2 to 3cm below the mark and shake for 10 to 15 minutes to completely dissolve the product. Make up to 100mL with deionized water, shake well, and then take 5mL of the solution and filter it through a 0.22μm organic filter membrane. Collect the filtrate. Detect the filtrate using a high-performance liquid chromatograph. Use an amino column, a mobile phase of acetonitrile and water at a volume ratio of 75:25, a flow rate of 0.8 to 1.2mL / min, a detection wavelength of 280nm, and an injection volume of 10μL.

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