Bile acid extraction waste treatment method and application

Abstract

This invention relates to the field of waste resource utilization and agricultural fertilizer technology. Specifically, it relates to a method and application for treating bile acid extraction waste. This method co-processes bile acid extraction waste with the products of harmless treatment of diseased and dead livestock and poultry into organic fertilizer, which can be used in tobacco cultivation. This invention proposes an integrated co-processing strategy of "two-stage enzymatic pretreatment - microbial fermentation - fertilization": a composite enzyme preparation is constructed by combining bromelain, acidic protease, keratinase, lipase, and xylanase; a two-stage process of acidic enzymatic hydrolysis and neutral main enzymatic hydrolysis is used to break down the complex organic structure of the two types of waste; ultrasonic assistance can be selected to enhance the degradation effect; then, a two-step high-temperature resistant composite microbial fermentation is carried out to achieve efficient degradation and transformation of organic matter; finally, functional components are added to optimize fertilizer efficiency. This invention achieves the co-resource utilization of two types of waste, and the prepared tobacco organic fertilizer has excellent growth-promoting and disease-resistant properties, combining environmental and economic benefits.

Description

Technical Field

[0001] This invention relates to the field of waste resource utilization and agricultural fertilizer technology. Specifically, it relates to a method and application for treating bile acid extraction waste. This method co-processes bile acid extraction waste with the products of harmless treatment of diseased and dead livestock and poultry into organic fertilizer, which can be used for tobacco planting and cultivation. Background Technology

[0002] The extraction of bile acids generates a large amount of waste, which is rich in fat and protein, typically exceeding 25% crude fat and 30% crude protein. However, because the fat encapsulates the protein in a stable structure, conventional treatment methods are inefficient in degrading it, resulting in extremely low resource utilization rates. This not only wastes resources but may also cause environmental pollution. Meanwhile, the harmless disposal of diseased and dead livestock and poultry often employs high-temperature, high-pressure sterilization processes. The resulting products contain large amounts of keratin and fibrous materials with a stubborn structure that is difficult for microorganisms to utilize, limiting their resource utilization pathways. Landfilling or incineration are the primary methods, wasting biomass resources and increasing the environmental burden.

[0003] Tobacco, as an important economic crop, has high requirements for soil fertility and fertilizer properties. Traditional organic fertilizers suffer from slow nutrient release and insufficient growth-promoting and disease-resistant properties, while organic fertilizers prepared from single wastes often have defects such as unbalanced nutrients and lack of functionality. Therefore, how to achieve efficient synergistic treatment of bile acid extraction waste and harmless treatment products of diseased and dead livestock and poultry to prepare high-quality organic fertilizer that meets the growth needs of tobacco has become an urgent technical problem to be solved. Summary of the Invention

[0004] To address the aforementioned technical problems, this invention discloses a method for treating bile acid extraction waste and its application, specifically including the following: I. Raw Material Selection and Proportioning The waste from bile acid extraction is the residue after bile acid extraction in industrial production, which is required to have a crude fat content of ≥25% and a crude protein content of ≥30%. The harmless treatment product of diseased and dead livestock and poultry is the pulverized product of diseased and dead pigs, chickens and other livestock and poultry after high temperature and high pressure sterilization at 121℃ and 0.1MPa for 2 hours, with a particle size of ≤5mm, a crude fiber content of ≥18%, and a keratin content of ≥10%. The two are mixed at a mass ratio of 1:2-3.

[0005] II. Multi-enzyme synergistic pretreatment system To address the high-fat, high-protein characteristics of bile acid extraction waste, lipase was selected to break down the lipid structure, and acidic protease was selected to degrade the protein. For the keratin and fiber structure of products from diseased and dead livestock and poultry, keratinase was selected. The enzymes were compounded at an enzyme activity ratio of 3:2:2:1, a ratio optimized through extensive experimentation to maximize enzymatic hydrolysis efficiency. The enzymatic hydrolysis conditions were controlled at pH 5.0-6.0 and temperature 45-50℃. This environment not only suits the optimal reaction conditions for various enzymes but also avoids the inhibition of the activity of a single enzyme. Through 4-6 hours of constant temperature stirring treatment, the degradation rate of crude fat in the mixed raw materials reached ≥60%, crude protein ≥50%, and crude fiber ≥40%, significantly improving the subsequent microbial utilization of nutrients.

[0006] III. High-Temperature Resistant Two-Step Compound Microbial Fermentation In the primary fermentation stage, a complex microbial community is constructed using actinomycetes, thermophilic fungi, and thermophilic Bacillus. Actinomycetes can degrade complex organic matter, thermophilic fungi can further decompose fibrous substances, and thermophilic Bacillus has strong enzyme production and organic matter degradation capabilities. The three are mixed at a colony count ratio of 2:1:3 and fermented at 55-65℃ for 7-10 days, which can rapidly degrade the residual organic matter after enzymatic hydrolysis and kill harmful microorganisms in the raw materials. In the secondary fermentation stage, a complex of humic bacteria, nitrogen-fixing bacteria, and phosphorus- and potassium-solubilizing bacteria is selected. Humic bacteria can promote humus formation, nitrogen-fixing bacteria can fix nitrogen from the air, and phosphorus- and potassium-solubilizing bacteria can convert insoluble phosphorus and potassium in the soil into available forms. Fermentation is carried out at 35-45℃ for 15-20 days to achieve deep transformation of organic matter and nutrient enrichment, thereby enhancing the fertilizer value of the fermentation products.

[0007] IV. Fertilizer Optimization and Characteristics of Tobacco Organic Fertilizer After the fermentation products are crushed and sieved, 3-5% humic acid and 1-2% seaweed extract are added. Humic acid can improve soil structure and increase nutrient utilization, while seaweed extract contains a variety of plant growth regulators, which can enhance the stress resistance and disease resistance of tobacco. The final prepared tobacco organic fertilizer has an organic matter content of ≥45%, total nutrients (N+P2O5+K2O) ≥5%, humic acid content ≥8%, and a pH of 6.5-7.5, which meets the soil pH and nutrient requirements for tobacco growth.

[0008] The beneficial effects of this invention are: (1) This invention addresses the structural problems of two types of waste through the innovative design of a multi-enzyme synergistic pretreatment system. Compared with single enzyme pretreatment, the degradation rate of organic matter is increased by more than 30%, laying a good foundation for subsequent fermentation. (2) The two-step fermentation of high temperature resistant compound microbial community realizes the synergistic effect of different functional microbial communities. The high temperature rapid degradation of the main fermentation is combined with the nutrient enrichment of the post fermentation, which shortens the fermentation cycle by 20-30% and makes the fermentation product more balanced in nutrients. (3) The integrated process of “enzyme pretreatment-microbial fermentation-fertilization” realizes the synergistic resource utilization of bile acid extraction waste and harmless treatment products of diseased and dead livestock and poultry. The utilization rate of the two types of waste reaches 100%, which solves the environmental problems of traditional treatment methods. (4) The prepared tobacco organic fertilizer has both growth-promoting and disease-resistant properties. When applied to tobacco planting, it can increase the tobacco seedling emergence rate by 10-15%, reduce the disease incidence rate by 20-25%, and increase the tobacco leaf yield by 15-20%, thus significantly improving the economic benefits of tobacco planting. Detailed Implementation

[0009] To enhance understanding of the present invention, the present invention will be further described in detail below with reference to embodiments. These embodiments are only used to explain the present invention and do not constitute a limitation on the scope of protection of the present invention.

[0010] Example 1: A method for treating waste from bile acid extraction (1) Raw material pretreatment: Dead livestock and poultry (such as pigs) are rendered harmless by high-temperature treatment (140℃, 40min) and mechanically crushed to obtain particles with a particle size of less than 5mm. The waste material after extracting deoxycholic acid from chicken bile is ground to form a uniform suspension. The two materials are mixed at a mass ratio (dead livestock and poultry product: bile extraction waste) = 1:0.2. Rice husks are added to adjust the carbon-nitrogen ratio (C / N) of the mixture to 30:1. Process water is added to adjust the moisture content of the mixture to 45%. The mixture is mechanically stirred at 75rpm for 30min in a mixing tank to form a uniform slurry. Calcium carbonate is added to adjust the pH of the mixture and maintain it at 5.5.

[0011] (2) Two-step enzymatic pretreatment: First stage (acidic enzymatic hydrolysis): Adjust the pH of the material to 3.0 and maintain the temperature at 37℃. Add acidic protease and react for 1.5 hours. The amount of acidic protease added is 20% of the total mass of the subsequent compound enzyme preparation.

[0012] Second stage (neutral main enzymatic hydrolysis): Adjust the pH of the material to 6.5 and raise the temperature to 52℃. Add a compound enzyme preparation consisting of 30 parts by weight of bromelain, 20 parts by weight of keratinase, 12 parts by weight of lipase, and 6 parts by weight of xylanase. The total amount of the compound enzyme preparation added is 1.0% of the mass of the mixed raw materials. React for 4 hours under slow stirring.

[0013] (3) High-temperature resistant two-step compound microbial fermentation: Microbial agent activation and addition: Main fermentation agent: freeze-dried powders of Streptomyces jingyangensis (ACCC40126), Chaetomium thermophilum (CGMCC3.14333), and Bacillus licheniformis (CGMCC1.10257) are mixed in a mass ratio of 1:0.5:0.5, activated with warm water (37℃) for 30 minutes, and then inoculated at 0.5% of the total mass of the material.

[0014] Post-fermentation inoculant: Mix the freeze-dried powders of Trichoderma harzianum (CGMCC3.10154), Azotobacter chrysotrichum (CGMCC1.16846), and Bacillus megaterium (CGMCC1.6721) in a mass ratio of 1:0.5:0.5, activate with warm water (37℃) for 30 minutes, and then set aside.

[0015] Primary fermentation (high-temperature stage): Place the inoculated material in a sealed fermenter, control the temperature at 55℃, maintain dissolved oxygen at 0.8 mg / L, and ferment for 6 days. During this period, turn the material twice a day with a slow stirrer (12 rpm).

[0016] Post-fermentation (cooling stage): After the main fermentation is completed, wait for the temperature to naturally drop to 42℃, then inoculate with the activated post-fermentation inoculum at a rate of 0.5% of the total material mass. Control the dissolved oxygen at 2.5 mg / L and continue fermentation for 3 days. Then allow the material to cool naturally to around 28℃ for 7 days of post-fermentation. Fermentation is considered complete when the germination index (GI) reaches 90%, the daily temperature difference is less than 1℃, and there is no ammonia odor.

[0017] (4) Preparation and application of tobacco organic fertilizer: The fully fermented material is vacuum-concentrated at low temperature to 1 / 3 of its original volume. 2% phosphate rock powder, 3% potassium feldspar, 2% boron, and 2% zinc (added in the form of trace element compounds) are added to the concentrated material and mixed evenly. After drying, crushing, and granulation, tobacco organic fertilizer is obtained. This organic fertilizer has an organic matter content ≥48%, total nutrients (N+P2O5+K2O) ≥5.5%, humic acid content ≥8%, and a pH value of 7.0. When applying, it is used as a base fertilizer and applied to the tobacco planting soil in one application at a rate of 2500 kg / hm². Example

[0018] (1) Raw material pretreatment: Take bile acid extraction waste with a crude fat content ≥25% and a crude protein content ≥30%, and mix it with the harmless treatment product of diseased and dead livestock and poultry (crude fiber ≥18%, keratin ≥10%) that has been treated at 121℃ and 0.1MPa for 2 hours and then crushed (particle size ≤5mm) at a mass ratio of 1:2.5. Adjust the moisture content of the mixture to 60% and set aside.

[0019] (2) Two-stage enzymatic pretreatment: Acidic enzymatic hydrolysis: Adjust the pH of the mixture to 3.0, maintain the temperature at 38℃, add an acidic protease with an enzyme activity ≥5000 U / g, and react for 1.5 hours. The amount of acidic protease added should account for 20% of the total mass of the compound enzyme preparation.

[0020] Neutral main enzymatic hydrolysis: Adjust the pH of the material to 6.8 and raise the temperature to 52℃. Add a compound enzyme preparation consisting of 30 parts of bromelain with an enzyme activity ≥6000U / g, 12 parts of keratinase with an enzyme activity ≥8000U / g, 8 parts of lipase with an enzyme activity ≥4000U / g, 7 parts of xylanase with an enzyme activity ≥10000U / g, and 20 parts of maltodextrin (by weight). The total addition amount is 1.0% of the mass of the mixed raw materials. React with slow stirring for 4 hours.

[0021] (Optional step): During the second stage of enzymatic hydrolysis, low-intensity ultrasonic treatment with a power of 0.4W / cm² and a frequency of 22kHz is used as an adjunct.

[0022] (3) High-temperature resistant two-step compound microbial fermentation: Primary fermentation: Inoculate the enzymatically hydrolyzed material with a primary fermentation agent (a mixture of actinomycetes, thermophilic Chaetomium, and Bacillus subtilis at a colony count ratio of 2:1:3), with an inoculation amount of 6% of the dry weight of the raw material. Control the fermentation temperature at 60℃ and ferment for 8 days, turning the material once a day.

[0023] Post-fermentation: After the main fermentation is completed, the temperature is lowered to 40℃, and a post-fermentation inoculum (a mixture of Aspergillus niger, Azotobacter chrysotile, and phosphorus- and potassium-solubilizing bacteria in a colony count ratio of 1:2:2) is inoculated at 4% of the dry weight of the raw material. Ferment at 40℃ for 18 days, turning the mixture every 2 days.

[0024] (4) Fertilizer optimization: The fermentation product was cooled to room temperature and pulverized through a 20-mesh sieve. 4% humic acid and 1.5% seaweed extract by weight of the fermentation product were added and mixed evenly to obtain tobacco organic fertilizer. The fertilizer has an organic matter content ≥50%, total nutrients ≥6.0%, humic acid content ≥10%, and pH 7.2. Example

[0025] (1) Raw material pretreatment: Bile acid extraction waste and products from the harmless treatment of diseased and dead livestock and poultry were mixed at a mass ratio of 1:3. This ratio resulted in a higher fibrous keratin content. More rice husk powder was added to adjust the C / N ratio to 35:1, the moisture content to 50%, and the pH to 5.8.

[0026] (2) Enzymatic pretreatment: Acidic enzymatic hydrolysis: pH 2.5, temperature 40℃, add acidic protease and react for 2 hours.

[0027] Neutral main enzymatic hydrolysis: pH 6.2, temperature 55℃. To address the high fibrous keratin properties, the ratio of keratinase and xylanase was appropriately increased. A complex enzyme consisting of 25 parts bromelain, 15 parts acidic protease, 15 parts keratinase, 10 parts lipase, and 8 parts xylanase was used, with a total addition amount of 1.2% of the raw material. The reaction time was 5 hours.

[0028] (3) Fermentation process: Primary fermentation: Inoculate with primary fermentation agents (Streptomyces jingyangensis, Aspergillus fumigatus CGMCC3.11251, and Bacillus pumilus CGMCC1.10291, mass ratio 1:1:1), inoculation amount 1.0%. Ferment at 58℃ for 7 days.

[0029] Post-fermentation: After cooling to 45℃, inoculate with post-fermentation inoculum (humic bacteria, nitrogen-fixing bacteria, and phosphorus- and potassium-solubilizing bacteria in a mass ratio of 1:1:1), with an inoculation amount of 1.0%. Maintain at 40-45℃ for 4 days, and then ripen for 10 days.

[0030] (4) Fertilizer preparation and effects: After fermentation, 3% humic acid and 1% seaweed extract were added, along with an additional 2% potassium feldspar to balance nutrients. The resulting organic fertilizer had an organic matter content ≥45%, total nutrients ≥5.0%, humic acid content ≥8%, and pH 6.8. In tobacco field trials, application at 2000 kg / hm² resulted in a 12% increase in tobacco emergence rate, a 22% decrease in mosaic virus incidence, and an 18% increase in tobacco yield compared to the control group.

[0031] The foregoing has shown and described the basic principles, main features, and advantages of the present invention. Those skilled in the art should understand that the present invention is not limited to the above embodiments. The embodiments and descriptions in the specification are merely illustrative of the principles of the invention. Various changes and modifications can be made to the invention without departing from its spirit and scope, and all such changes and modifications fall within the scope of the present invention as claimed. The scope of protection of the present invention is defined by the appended claims and their equivalents.

Claims

1. A method for processing waste of bile acid extraction, characterized by, Includes the following steps: Step 1: Raw material pretreatment. Mix the waste from bile acid extraction with the harmless treatment product of diseased and dead livestock and poultry at a mass ratio of 1:2-3, and adjust the moisture content of the mixture to 55-65% for later use. Step 2, two-stage enzymatic pretreatment, includes the following process: Process 1, acidic enzymatic hydrolysis: Adjust the pH of the mixed raw materials in step 1 to 2.0-3.5, maintain the temperature at 35-40℃, add acidic protease and react for 1-2 hours; Step 2, Neutral main enzymatic hydrolysis: Adjust the pH of the material treated in Step 1 to 6.0-7.0, raise the temperature to 50-55℃, add the compound enzyme preparation, and stir slowly for 3-5 hours. The total amount of compound enzyme preparation added is 0.8-1.2% of the mass of the mixed raw materials. Step 3: High-temperature resistant two-step compound microbial fermentation, including the following process: Process 1, Primary Fermentation: Inoculate the pretreated raw materials from Step 2 with the primary fermentation agent, adjust the temperature to 55-65℃, and ferment for 7-10 days, turning and stirring 1-2 times daily during this period. The primary fermentation agent is a mixture of actinomycetes, thermophilic fungi, and thermophilic Bacillus at a colony count ratio of 2:1:3, and the inoculation amount is 5-8% of the raw material mass. Step 2, Post-fermentation: After the main fermentation is completed, the temperature is lowered to 35-45℃, and a post-fermentation inoculum is inoculated. Fermentation lasts for 15-20 days, during which the bacteria are turned over once every 2 days. The post-fermentation inoculum is a mixture of humic bacteria, nitrogen-fixing bacteria, and phosphorus-solubilizing and potassium-solubilizing bacteria in a colony count ratio of 1:2:

2. The inoculation amount is 3-5% of the raw material mass. Step 4, Fertilizer Optimization: After cooling the fermentation product from Step 3 to room temperature, crush it through a 20-mesh sieve, add humic acid and seaweed extract, mix well, and the basic organic fertilizer is obtained.

2. The method for treating waste of bile acid extraction according to claim 1, characterized by, The crude fat content of the bile acid extraction waste in step one is ≥25% and the crude protein content is ≥30%. The harmless treatment product of diseased and dead livestock and poultry is the pulverized product of diseased and dead livestock and poultry after high temperature and high pressure sterilization treatment. Its crude fiber content is ≥18% and keratin content is ≥10%, and it contains hair and hoof keratin components.

3. The method according to claim 2, wherein In step two, the amount of acidic protease added is 15-25% of the total mass of the compound enzyme preparation. The compound enzyme preparation consists of 25-35 parts of bromelain, 10-15 parts of keratinase, 5-10 parts of lipase, and 5-8 parts of xylanase by weight. The enzyme activity of bromelain is ≥6000 U / g, the enzyme activity of acidic protease is ≥5000 U / g, the enzyme activity of keratinase is ≥8000 U / g, the enzyme activity of lipase is ≥4000 U / g, and the enzyme activity of xylanase is ≥10000 U / g.

4. The method according to claim 3, wherein In step two, during the neutral enzymatic hydrolysis process of process 2, low-intensity ultrasonic treatment is used, with an ultrasonic power of 0.3-0.5 W / cm² and a frequency of 20-25kHz.

5. The method according to claim 4, wherein the waste containing bile acids is waste containing a large amount of bile acids. The thermophilic fungi mentioned in step three are thermophilic Chaetomium or thermophilic Ascomycetes, and the thermophilic Bacillus is thermophilic subsp. subsp. subtilis or thermophilic variant of Bacillus licheniformis.

6. The method according to claim 5, wherein the waste containing bile acids is waste containing a large amount of bile acids. In step four, the amount of humic acid added is 3-5% of the mass of the fermentation product, and the amount of seaweed extract added is 1-2% of the mass of the fermentation product.

7. An organic fertilizer for tobacco cultivation, characterized by, The basic organic fertilizer obtained by treatment as described in claim 6 has an organic matter content of ≥48%, a total nutrient content of N+P2O5+K2O ≥5.5%, a humic acid content of ≥8%, and a pH of 6.5-7.

5. The application rate of the organic fertilizer is 2000-3000 kg / hm², and it is applied to the planting soil as a base fertilizer in one application.