A production process of enzymatic bone powder rich in low-molecular sodium chondroitin sulfate

Abstract

The invention provides a production process of enzymatic bone powder rich in low-molecular sodium chondroitin sulfate, belonging to the preparation technology of biomedical raw materials and food raw materials. The method is characterized in that the ceramic membrane filtration interception liquid in the process is circulated and treated, and continuous countercurrent extraction and separation are adopted to continuously separate the oil and fat and low-molecular-weight polypeptides, mucopolysaccharides and other substances in the hydrolyzate, reduce the concentration of the hydrolyzate, increase the enzymatic hydrolysis speed, and control the degree of hydrolysis, thereby improving the product yield within the molecular weight control range of the hydrolyzate, and at the same time prevent the accumulation of oil and fat from bonding bone particles to affect the release of soluble substances in the bone particles, increase the adsorption amount of protease and bone particles, improve the hydrolysis speed and the yield of enzymatic bone powder, and the method can also co-produce collagen peptides, sodium chondroitin sulfate, amino acid powder, tallow and high calcium powder, so as to realize clean production with full utilization of raw materials and reduced discharge of three wastes.

Description

Technical Field

[0001] The invention relates to a preparation technology of biomedical raw materials and food raw materials, in particular to a production process of enzymatic bone powder rich in low-molecular sodium chondroitin sulfate. Background Art

[0002] Sodium chondroitin sulfate (CS for short) is an acidic mucopolysaccharide, which is widely found in cattle bones, pig bones, chicken bones, shark bones, and squid bones. It is a type of glycosaminoglycan and a natural biological macromolecule. Sodium chondroitin sulfate is composed of 50-70 disaccharide units, which are formed by glucuronic acid and aminogalactose through β-1,3 glycosidic bonds. Disaccharides are then connected and polymerized into macromolecules with β-1,4 glycosidic bonds. It has physiological functions such as lowering blood lipids, anti-coagulation, anti-inflammation and anti-tumor. It is mainly used clinically for the treatment of osteoarthritis, hyperlipidemia, angina pectoris, atherosclerosis and other diseases. Its eye drops can treat corneal damage, corneal ulcers or keratitis. In food, sodium chondroitin sulfate is classified as a dietary supplement by the US FDA, and Australia regards it as a nutritional health product for improving or preventing joint diseases.

[0003] At present, sodium chondroitin sulfate is extracted from animal cartilage by enzymatic hydrolysis. Animal cartilage, especially bovine cartilage, is a very high-quality material that is easy to obtain in large quantities for industrial production. It is rich in valuable ingredients such as chondroitin sulfate, collagen, oil, and calcium. In the prior art, a large amount of usable material is discharged during the extraction of sodium chondroitin sulfate from bovine cartilage, and the yield of sodium chondroitin sulfate is not ideal. How to improve the yield of sodium chondroitin sulfate, and simultaneously produce collagen, oil, amino acid powder and high calcium powder, so as to achieve clean production with full utilization of bovine cartilage and reduced discharge of three wastes, is an issue that needs to be solved and improved urgently. Summary of the invention

[0004] In order to make up for the deficiencies in the prior art, the present invention provides a new process for the co-production of enzymatic bone powder rich in sodium chondroitin sulfate based on animal cartilage, which achieves an ideal enzymatic bone powder yield and reduces heavy metal residues in the product; collagen peptides, sodium chondroitin sulfate, amino acid powder, oil and high calcium powder can be co-produced at the same time, cattle bones can be fully utilized, the discharge of three wastes is reduced, and clean production is achieved.

[0005] The first aspect of the present invention provides a production process of enzymatic bone powder rich in low molecular weight sodium chondroitin sulfate, characterized in that it comprises the following steps:

[0006] S1. Pretreatment: The animal cartilage is steamed, and then the floating oil is removed, and then the animal cartilage is crushed in an extraction tank; the extraction tank is provided with a stirring paddle and a baffle with a shearing function at the bottom, the tank body is in a downward cone shape, and the bottom cover has a water inlet pipe and a valve at the bottom; the animal cartilage is crushed into fine particles by shearing with a stirring paddle with a shearing function and squeezing between bone particles during the stirring process, and the overall liquid is in a slurry state;

[0007] S2. Enzymatic hydrolysis: Cool the liquid to 50-55 degrees, adjust the pH to 7-8, and then add protease for 0.5-2 hours.

[0008] S3. Degreasing by adsorption: the enzymatic solution is released from the upper part of the extraction tank to the degreasing tank for degreasing by adsorption;

[0009] S4. Ceramic membrane filtration: The deoiled liquid is collected in the first extract transfer tank and then pumped into the ceramic membrane filter for filtration;

[0010] S5. Circulate the retentate filtered by the ceramic membrane: reflux the retentate filtered by the ceramic membrane into the second extract transfer tank, that is, the feed liquid in the second extract transfer tank is introduced into the extract tank through the bottom water inlet pipe of the extract tank to perform the enzymatic hydrolysis in step S2; S3 the enzymatic hydrolyzate passes through the bone granular layer in the extract tank and is released from the top to flow into the deoiling tank; S4 the feed liquid after adsorption and deoiling is collected in the first extract transfer tank, and then pumped into the ceramic membrane filter for filtration, and the generated retentate filtered by the ceramic membrane continues to circulate and reflux to the second extract transfer tank;

[0011] The circulation process is continued until there are no obvious cartilage particles in the liquid material in the second extraction liquid transfer tank, and the circulation is terminated when the liquid material is clear;

[0012] During the circulation process, reverse osmosis water is added to the second extract transfer tank to make up for the water lost by the ceramic membrane filtration;

[0013] S6. After the circulation treatment is completed, the liquid in the second extraction liquid transfer tank is pumped back to the extraction tank through the reflux pump, and the stirring speed of the extraction tank is increased. After 20-80 minutes, the liquid in the extraction tank is pumped into the self-unloading flat plate centrifuge, and the filtrate is collected through the extraction transfer tank, and then added to the cation exchange resin tank through the carbon removal filter. After soaking for 2-5 hours, it is pumped into the ultrafiltration liquid transfer tank for subsequent ultrafiltration using an ultrafiltration unit. The ultrafiltration retentate is an enzymatic bone powder aqueous solution.

[0014] The preferred embodiment of the production process of the present invention further comprises preparing enzymatic bone powder based on the ultrafiltration retentate produced by S6 by one or both of the following methods:

[0015] (1) concentrating the ultrafiltration retentate to a water content of 50-70%, and then spray-drying to obtain enzymatic bone powder;

[0016] (2) After the ultrafiltration retentate is concentrated to 10%, reverse osmosis water is added to concentrate it to 10% again, and then alcohol precipitation, dehydration, granulation, drying, crushing and sieving are performed to obtain enzymatic bone powder.

[0017] In the preferred embodiment of the production process of the present invention, the protease in S2 is a mixed enzyme of 10000U / g flavor protease and 200000U / g alkaline protease in a mass ratio of 2:1, and the addition ratio is 0.5-0.9% of the mass of the feed liquid.

[0018] Another aspect of the present invention provides a production process for the co-production of collagen peptides, sodium chondroitin sulfate and amino acid powder based on animal cartilage, characterized in that it comprises the following steps:

[0019] S1. Pretreatment: The animal cartilage is steamed, and then the floating oil is removed, and then the animal cartilage is crushed in an extraction tank; the extraction tank is provided with a stirring paddle and a baffle with a shearing function at the bottom, the tank body is in a downward cone shape, and the bottom cover has a water inlet pipe and a valve at the bottom; the animal cartilage is crushed into fine particles by shearing with a stirring paddle with a shearing function and squeezing between bone particles during the stirring process, and the overall liquid is in a slurry state;

[0020] S2. Enzymatic hydrolysis: cool the liquid to 50-55 degrees, adjust the pH to 7-8, and then add protease for 0.5-2 hours;

[0021] S3 adsorption and degreasing: the enzymatic hydrolyzate is discharged from the upper part of the extraction tank to the degreasing tank for adsorption and degreasing;

[0022] S4. Ceramic membrane filtration: The deoiled feed liquid is collected in the first extract transfer tank, and then pumped into the ceramic membrane filter for filtration; S4.1 The ceramic membrane filtered permeate is pumped into a nanofiltration concentration unit with a 300 Dalton nanofiltration membrane for nanofiltration, and after concentrating to 15%, 1 volume of reverse osmosis water is added, and then concentrated again, and the nanofiltration retentate is temporarily stored in the nanofiltration retentate transfer tank, and then vacuum concentrated and dried to obtain bovine collagen peptides; the nanofiltration permeate is stored in a storage tank;

[0023] S4.2 The retentate filtered by the ceramic membrane is circulated: the retentate filtered by the ceramic membrane is refluxed and introduced into the second extract transfer tank, that is, the feed liquid in the second extract transfer tank is introduced into the extraction tank through the bottom water inlet pipe of the extraction tank to perform the enzymatic hydrolysis in step S2; S3 The enzymatic hydrolyzate passes through the bone granular layer in the extraction tank and is released from the top to flow into the deoiling tank; S4 The feed liquid after adsorption and deoiling is collected in the first extract transfer tank, and then pumped into the ceramic membrane filter for filtration.

[0024] The ceramic membrane filtration retentate produced continues to circulate back to the second extraction liquid transfer tank; the ceramic membrane filtration permeate produced in the circulation treatment is treated in the manner of S4.1 to produce nanofiltration permeate;

[0025] The circulation process is continued until there are no obvious cartilage particles in the liquid material in the second extraction liquid transfer tank, and the circulation is terminated when the liquid material is clear;

[0026] During the circulation process, reverse osmosis water is added to the second extract transfer tank to make up for the water lost by the ceramic membrane filtration;

[0027] S5. The second extract transfer tank liquid is pumped back to the extraction tank through the reflux pump, the stirring speed of the extraction tank is increased, and the liquid in the extraction tank is pumped into the self-unloading flat centrifuge after 20-80 minutes, and the filtrate is collected by the extraction transfer tank, and then added to the cation exchange resin tank through the carbon removal filter, and then soaked for 2-5 hours and pumped into the ultrafiltrate transfer tank for subsequent ultrafiltration using an ultrafiltration unit, and the ultrafiltration retentate is an enzymatic bone meal aqueous solution;

[0028] S6. The ultrafiltration permeate produced in S5 is treated in the same manner as in S4.1 to produce a nanofiltration permeate;

[0029] S7. The nanofiltration permeate produced in steps S4 and S6 is transferred to a reverse osmosis transfer tank for reverse osmosis. The reverse osmosis retentate is concentrated to 25% and then vacuum concentrated to a solid content of more than 40%, and then spray-dried to obtain amino acid powder.

[0030] In the preferred embodiment of the above-mentioned co-production process, the protease in S2 is a mixed enzyme of 10000U / g flavor protease and 200000U / g alkaline protease in a mass ratio of 2:1, and the addition ratio is 0.5-0.9% of the mass of the feed liquid.

[0031] In the preferred embodiment of the above-mentioned co-production process, the vacuum concentration and drying of the nanofiltration retentate described in step 4.1 is first vacuum concentrated to a solid content of 50%, and then transferred to a spray drying transfer tank, and the drying temperature is controlled to an inlet air temperature of 170-190 degrees Celsius and an outlet air temperature of 80-90 degrees Celsius.

[0032] In the preferred embodiment of the above-mentioned co-production process, the filter residue produced by centrifugation in step S5 is dried and crushed to obtain high calcium powder.

[0033] The preferred embodiment of the above-mentioned co-production process further includes collecting the oil and fat obtained by degreasing in steps S1, S3 and S4.

[0034] In the above process, the animal cartilage includes one or more of bovine cartilage, chicken cartilage, fish cartilage, and pig cartilage. Bovine cartilage can be nasal bones, trachea, rib cartilage, and shoulder blades; chicken cartilage comes from chicken leg bones; fish cartilage can be shark cartilage or other fish cartilage; and pig cartilage can be trachea. In the third aspect of the present invention, a method for extracting a can in the above production process is also provided.

[0035] An extraction tank for producing enzymatic bone powder rich in low-molecular sodium chondroitin sulfate, characterized in that: it has a cylindrical tank body 1, the lower part of the tank body 1 is conical; a motor 2 is arranged on the top of the tank body, a stirring shaft 3 controlled by the motor 2 is located at the axis position of the tank body and extends to the bottom of the cone, a stirring paddle 4 with a shearing function is arranged at the lower part of the stirring shaft 2, and stirring paddles 5 are arranged in the middle and upper parts;

[0036] The bottom cover of the tank body is provided with a bottom water inlet pipe 6, a valve 7, and an air inlet pipe 8; the top of the tank body is provided with a discharge port 9 for discharging the enzymatic hydrolysis liquid into the oil removal tank; a feed hole 10, an alkali pipe 11, and an acid pipe 12 are also provided;

[0037] In some preferred embodiments, a lower temperature detection tube 13, a middle temperature detection tube 14 and an upper temperature detection tube 15 are sequentially arranged on the side wall of the tank body 1 from bottom to top;

[0038] In some preferred embodiments, a sampling and analysis tube 16 and a disinfectant tube 17 are also provided on the side wall of the tank body 1 .

[0039] During use, the stirring paddle 4 with a shearing function cuts and squeezes the bone particles during the stirring process to crush the animal cartilage into fine particles, and the entire liquid feed is in a slurry state.

[0040] The present invention also provides an equipment set for producing enzymatic bone powder rich in low-molecular sodium chondroitin sulfate, characterized in that it comprises any of the above-mentioned extraction tanks, an oil removal tank, a first extraction liquid transfer tank, a ceramic membrane filter and a second extraction liquid transfer tank connected by a pipeline closed loop, and is used for cyclic treatment of animal cartilage;

[0041] The extraction tank is used to stir, crush and hydrolyze the animal cartilage with enzymes;

[0042] The liquid inlet of the de-oiling tank is connected to the discharge port 9 provided at the upper part of the extraction tank through a pipeline, so that the enzymatic hydrolysis liquid in the extraction tank can be discharged into the de-oiling tank for de-oiling;

[0043] The first extraction liquid transfer tank is used to collect the deoiled liquid from the deoiling tank.

[0044] The ceramic membrane filter is used to filter the liquid from the first extraction liquid transfer tank.

[0045] The second extraction liquid transfer tank is used to collect the filtered intercepted liquid produced by the ceramic membrane filter. It is connected to the bottom water inlet pipe 6 at the bottom of the extraction tank through a pipeline, so that the filtered intercepted liquid is returned to the extraction tank through the bottom water inlet pipe 6 through a reflux pump, thereby realizing the circulation treatment of the feed liquid.

[0046] In a preferred embodiment, the equipment combination also includes an ultrafiltration transfer tank and an ultrafiltration device. The ultrafiltration transfer tank is used to collect the liquid that has completed the circulation treatment in the extraction tank, and the ultrafiltration device is used to ultrafilter the liquid that has completed the circulation treatment. The intercepted liquid produced by the ultrafiltration is post-treated to obtain enzymatic bone powder.

[0047] Based on the above process description of the present invention, the equipment combination is assembled with conventional nanofiltration membrane concentration units, nanofiltration devices, reverse osmosis equipment, thermal concentration equipment, and various containers for liquid transfer in the art to obtain an equipment group for co-producing collagen peptides, sodium chondroitin sulfate and amino acid powder ( Figure 1 shown).

[0048] Beneficial effects of the present invention:

[0049] The production process for producing enzymatic bone powder based on cartilage provided by the present invention realizes shearing, grinding, fluid flushing and dissolution, and solid-liquid separation in an extraction tank, can continuously crush cartilage and release water-soluble substances such as collagen and glycoprotein, the concentration of the dissolved matter is controllable, and the production capacity of a single tank can be increased.

[0050] In the process, the ceramic membrane filtration retentate is circulated and treated, and continuous countercurrent extraction and separation are adopted to continuously separate the oil and low molecular weight polypeptides, mucopolysaccharides and other substances in the hydrolyzate, thereby reducing the concentration of the hydrolyzate, increasing the enzymatic hydrolysis rate, and controlling the degree of hydrolysis, thereby increasing the product yield within the molecular weight control range of the hydrolyzate. At the same time, it can prevent the accumulation of oil and fat from adhering to the bone particles and affecting the release of soluble substances in the bone particles, increase the adsorption amount of protease and bone particles, increase the hydrolysis rate, and at the same time improve the yield of enzymatic bone powder. No chemical extractant is used in the process.

[0051] Experimental data show that, taking bovine cartilage as raw material, the yield of enzymatic bone powder is 6%-10%, of which sodium chondroitin sulfate (of which 30-50% has a molecular weight of less than 5000) accounts for 90-95%, protein 2-4%, and sodium hyaluronate 0.5-1%.

[0052] During the whole process, the pH is controlled at 7-8, which prevents the dissolution of calcium, magnesium and other minerals in the bones, increases the adsorption of cation exchange resin to remove metal ions, reduces the ash content, greatly reduces the residual heavy metal ions, and improves the quality of enzymatic bone powder.

[0053] A 300-dalton nanofiltration membrane is used for concentration to remove small molecules with a molecular weight below 300, such as chloride ions, sulfate ions, amino acids, etc., thereby increasing the peptide content of collagen peptides.

[0054] In the co-production process, enzymatic bone meal and collagen peptides are concentrated separately, which can significantly shorten the production cycle and separately concentrate chondroitin and peptides to reduce product discoloration.

[0055] It can co-produce enzymatic bone powder (sodium chondroitin sulfate), collagen peptides, amino acid powder, high calcium powder and oil, fully utilize the cartilage, significantly reduce the discharge of three wastes, and achieve clean production. BRIEF DESCRIPTION OF THE DRAWINGS

[0056] Figure 1 A schematic diagram of the production process of the present invention for co-producing animal cartilage to prepare collagen peptides, sodium chondroitin sulfate and amino acid powder;

[0057] Figure 2 A schematic diagram of the structure of the extraction tank used in the present invention;

[0058] Figure 3 . A schematic diagram of the connection of the equipment group used in the production process of the present invention for producing enzymatic bone powder rich in sodium chondroitin sulfate;

[0059] Among them, 1-tank body 1, 2-motor 2, 3-stirring shaft, 4-stirring paddle 4 with shearing function, 5-stirring paddle; 6-bottom water inlet pipe, 7-valve, 8-air inlet pipe, 9-discharge port, 10-feeding hole, 11-alkali pipe, 12-acid pipe, 13-lower temperature detection pipe, 14-middle temperature detection pipe, 15-upper temperature detection pipe; 16-sampling and analysis pipe, 17 and disinfectant pipe;

[0060] 21- oil removal tank, 22- first extraction liquid transfer tank, 23- ceramic membrane filter, 24- second extraction liquid transfer tank; DETAILED DESCRIPTION

[0061] The specific implementation process of the present invention is described below in conjunction with the accompanying drawings. The following description is only used to illustrate the present invention and is not used to limit the scope of protection of the present invention. Unless otherwise defined, the professional terms used in the present invention have the same meanings as those generally understood by those skilled in the art.

[0062] like Figure 2 As shown, the extraction tank used in the present invention has a cylindrical tank body 1, and the lower part of the tank body 1 is conical;

[0063] A motor 2 is arranged on the top of the tank body, and a stirring shaft 3 controlled by the motor 2 is located at the axis center of the tank body and extends to the conical bottom. A stirring paddle 4 with a shearing function is arranged at the lower part of the stirring shaft 2, and stirring paddles 5 are arranged in the middle and upper parts;

[0064] The bottom cover of the tank body is provided with a bottom water inlet pipe 6, a valve 7, and an air inlet pipe 8; the top of the tank body is provided with a discharge port 9 for discharging the enzymatic hydrolysis liquid into the oil removal tank; a feed hole 10, an alkali pipe 11, and an acid pipe 12 are also provided;

[0065] In some preferred embodiments, a lower temperature detection tube 13, a middle temperature detection tube 14 and an upper temperature detection tube 15 are sequentially arranged on the side wall of the tank body 1 from bottom to top;

[0066] In some preferred embodiments, a sampling and analysis tube 16 and a disinfectant tube 17 are also provided on the side wall of the tank body 1 .

[0067] During use, the stirring paddle 4 with a shearing function cuts and squeezes the bone particles during the stirring process to crush the bovine cartilage into fine particles, and the entire liquid feed is in a slurry state.

[0068] The present invention also provides an equipment group for producing enzymatic bone meal, such as Figure 3 As shown, it includes any of the above extraction tanks connected by a pipeline closed loop, an oil removal tank 21, a first extraction liquid transfer tank 22, a ceramic membrane filter 23 and a second extraction liquid transfer tank 24, which are used for cyclic processing of animal cartilage;

[0069] The extraction tank is used to stir, crush and hydrolyze the animal cartilage with enzymes;

[0070] The liquid inlet of the de-oiling tank is connected to the discharge port 9 provided at the upper part of the extraction tank through a pipeline, so that the enzymatic hydrolysis liquid in the extraction tank can be discharged into the de-oiling tank for de-oiling;

[0071] The first extraction liquid transfer tank is used to collect the deoiled liquid from the deoiling tank.

[0072] The ceramic membrane filter is used to filter the liquid from the first extraction liquid transfer tank.

[0073] The second extraction liquid transfer tank is used to collect the filtered intercepted liquid produced by the ceramic membrane filter. It is connected to the bottom water inlet pipe 6 at the bottom of the extraction tank through a pipeline, so that the filtered intercepted liquid is returned to the extraction tank through the bottom water inlet pipe 6 through a reflux pump, thereby realizing the circulation treatment of the feed liquid.

[0074] The equipment combination also includes an ultrafiltration transfer tank and an ultrafiltration device. The ultrafiltration transfer tank is used to collect the slurry that has completed the circulation treatment in the extraction tank, and the ultrafiltration device is used to ultrafilter the slurry that has completed the circulation treatment. The intercepted liquid produced by the ultrafiltration is post-treated to obtain enzymatic bone powder.

[0075] Example 1 Preparation of enzymatic bone powder rich in sodium chondroitin sulfate

[0076] Materials and reagents

[0077] Animal cartilage: cow nose bone

[0078] Compound protease: a mixed enzyme of 10000U / g flavor protease and 200000U / g alkaline protease at a mass ratio of 2: 1. Flavor protease and alkaline protease were purchased from Shandong Longkote Enzyme Preparation Co., Ltd.

[0079] Experimental setup

[0080] Extraction tank: Figure 2 As shown: extraction tank 30 cubic meters, oil removal tank 10 cubic meters, oil absorption fiber hanging inside,

[0081] The extraction liquid transfer tank is 10 cubic meters and was purchased from Yiyang Non-standard Equipment Factory.

[0082] Ceramic membrane filter, 3000 Dalton, 30 m3 / h, purchased from Hunan Henghui Membrane Technology Co., Ltd.;

[0083] Nanofiltration membrane concentrator unit, nanofiltration membrane, NF400, 8040 type, purchased from Zibo Huibang Environmental Engineering Co., Ltd.;

[0084] Cation exchange resin tank 10 cubic meters, resin, Rohm and Haas, purchased from Yiyang Non-standard Equipment Factory;

[0085] Ultrafiltration unit, ultrafiltration membrane SS-UE003-8040, purchased from Zibo Huibang Environmental Engineering Co., Ltd.;

[0086] Reverse osmosis water production device: RO-JS2-2000 Huaian Jinnuo Pharmaceutical Equipment Co., Ltd.

[0087] In the process of the present invention, except for the specially made extraction tank, the other required equipment are all commonly used equipment in the field, and those skilled in the art can replace them as needed.

[0088] The process steps are as follows, and the connection relationship and process of the main devices used are as follows: Figure 1 shown.

[0089] S1. Pretreatment: Remove meat, fat and other impurities from animal cartilage (fresh beef tendon), add 6 times the volume of water, heat to 105 degrees and steam for 7 hours, cool to 95 degrees, remove the floating oil; add to the extraction tank for stirring and crushing, stir for 1.5 hours, use shearing and squeezing between bone particles to crush the animal cartilage into coarse sand, and the liquid is slightly slurry;

[0090] S2. Enzymatic hydrolysis: The liquid is cooled to 50-55 degrees, the pH value is adjusted to 7-8, and then 0.9% of the mass of the liquid is added with a composite protease for enzymatic hydrolysis for 1 hour, and then transferred to the extraction reflux transfer tank;

[0091] S3 adsorption and degreasing: the enzymatic hydrolyzate is discharged from the upper part of the extraction tank to the degreasing tank for adsorption and degreasing;

[0092] S4. Ceramic membrane filtration: The deoiled feed liquid is collected in the first extract transfer tank, and then pumped into a 3000 Dalton ceramic membrane filter for filtration; S4.1 The ceramic membrane filtered permeate is pumped into a nanofiltration concentration unit with a 300 Dalton nanofiltration membrane for nanofiltration, and after concentrating to 15%, 1 volume of reverse osmosis water is added, and then concentrated again, and the nanofiltration retentate is temporarily stored in the nanofiltration retentate transfer tank, and then vacuum concentrated and dried to obtain bovine collagen peptides; the nanofiltration permeate is stored in a storage tank;

[0093] S4.2 Circulation treatment of the ceramic membrane filtration retentate: the ceramic membrane filtration retentate is refluxed and introduced into the second extraction liquid transfer tank, that is, the feed liquid in the second extraction liquid transfer tank is introduced into the extraction tank through the bottom water inlet pipe of the extraction tank to perform the enzymatic hydrolysis in step S2; S3 the enzymatic hydrolyzate passes through the bone granular layer in the extraction tank and is released from the top to flow into the deoiling tank; S4 the feed liquid after adsorption and deoiling is collected in the first extraction liquid transfer tank, and then pumped into a 3000 Dalton ceramic membrane filter for filtration, and the generated ceramic membrane filtration retentate continues to circulate and reflux to the second extraction liquid transfer tank; the ceramic membrane filtration permeate generated in the circulation treatment is treated in the manner of S4.1 to generate nanofiltration permeate;

[0094] The cycle is continued for 8 hours, or until there are no obvious cartilage particles in the liquid in the second extraction liquid transfer tank and the liquid is clear, and the cycle is terminated;

[0095] During the circulation process, reverse osmosis water is added to the second extract transfer tank to make up for the water lost by the ceramic membrane filtration;

[0096] S5. The second extract transfer tank liquid is pumped back to the extraction tank through the reflux pump, the stirring speed of the extraction tank is increased, and the liquid in the extraction tank is pumped into the self-unloading flat centrifuge after 60 minutes, and the filtrate is collected by the extraction transfer tank, and then the cation exchange resin tank is added through the carbon removal filter, and the ultrafiltrate is pumped into the ultrafiltration transfer tank after soaking for 5 hours for subsequent ultrafiltration using an ultrafiltration unit, and the ultrafiltration retentate is an enzymatic bone meal aqueous solution;

[0097] S6. The ultrafiltration permeate produced in S5 is treated in the manner of S4.1 to produce a nanofiltration permeate and a nanofiltration retentate;

[0098] S7. The nanofiltration permeate produced in steps S4 and S6 is transferred to a reverse osmosis transfer tank for reverse osmosis. The reverse osmosis retentate is concentrated to a solid content of 25% and then transferred to a nanofiltration retentate transfer tank. The mixture is then vacuum concentrated to a solid content of more than 40% and then spray dried to obtain amino acid powder.

[0099] S8. The nanofiltration retentate produced by the treatment in S4.1 is first vacuum concentrated (temperature 45-55°C) to a solid content of 50%, and then transferred to a spray drying transfer tank. The spray drying inlet air temperature is 170-190°C, and the outlet air temperature is 80-90°C to obtain bovine collagen peptides.

[0100] S9. The ultrafiltration retentate produced by S5 is concentrated to a water content of 50-70%, and then spray dried (spray drying inlet air temperature 170-190 degrees, outlet air 80-90 degrees) to obtain enzymatic bone powder;

[0101] Or the ultrafiltration retentate is concentrated to a solid content of 10%, and then reverse osmosis water is added to 1 times to concentrate it to a solid content of 10%, and then 60-65% ethanol is used for alcohol precipitation, the precipitate is dehydrated with alcohol, granulated, dried at 75 degrees Celsius, crushed, and sieved through an 80-mesh sieve to obtain enzymatic bone powder.

[0102] Example 2.

[0103] Materials and equipment are the same as in Example 1

[0104] S1. Pretreatment: Remove meat, fat and other impurities from animal cartilage (cow nose bone), add 8 times the volume of water, heat to 105 degrees and steam for 8 hours, cool to 95 degrees, remove the floating oil; add to the extraction tank for stirring and crushing, stir for 1.5 hours, use shearing and extrusion between bone particles to crush the animal cartilage into coarse sand, and the liquid is slightly slurry;

[0105] S2. Enzymatic hydrolysis: The liquid is cooled to 50-55 degrees, the pH value is adjusted to 7-8, and then a composite protease is added at a ratio of 0.5% of the liquid mass for 1.5 hours, and then transferred to the extraction reflux transfer tank;

[0106] S3 adsorption and degreasing: the enzymatic hydrolyzate is discharged from the upper part of the extraction tank to the degreasing tank for adsorption and degreasing;

[0107] S4. Ceramic membrane filtration: The deoiled feed liquid is collected in the first extract transfer tank, and then pumped into a 3000 Dalton ceramic membrane filter for filtration; S4.1 The ceramic membrane filtered permeate is pumped into a nanofiltration concentration unit with a 300 Dalton nanofiltration membrane for nanofiltration, and after concentrating to 15%, 1 volume of reverse osmosis water is added, and then concentrated again, and the nanofiltration retentate is temporarily stored in the nanofiltration retentate transfer tank, and then vacuum concentrated and dried to obtain bovine collagen peptides; the nanofiltration permeate is stored in a storage tank;

[0108] S4.2 Circulation treatment of the ceramic membrane filtration retentate: the ceramic membrane filtration retentate is refluxed and introduced into the second extraction liquid transfer tank, that is, the feed liquid in the second extraction liquid transfer tank is introduced into the extraction tank through the bottom water inlet pipe of the extraction tank to perform the enzymatic hydrolysis in step S2; S3 the enzymatic hydrolyzate passes through the bone granular layer in the extraction tank and is released from the top to flow into the deoiling tank; S4 the feed liquid after adsorption and deoiling is collected in the first extraction liquid transfer tank, and then pumped into a 3000 Dalton ceramic membrane filter for filtration, and the generated ceramic membrane filtration retentate continues to circulate and reflux to the second extraction liquid transfer tank; the ceramic membrane filtration permeate generated in the circulation treatment is treated in the manner of S4.1 to generate nanofiltration permeate;

[0109] The circulation treatment is continued for 7 hours until there are no obvious cartilage particles in the liquid material in the second extraction liquid transfer tank and the circulation is terminated after the liquid material becomes clear; during the circulation process, reverse osmosis water is added to the extraction reflux transfer tank to supplement the water lost by the ceramic membrane filtration;

[0110] S5. After the cycle is completed, the second extract transfer tank is fully pumped back to the extraction tank by a reflux pump to increase the stirring speed of the extraction tank. After 45 minutes, the extract tank is pumped into a self-unloading flat centrifuge, and the filtrate is collected by the extraction transfer tank, and then added to the cation exchange resin tank through a carbon removal filter, and soaked for 2 hours and pumped into the ultrafiltrate transfer tank for subsequent ultrafiltration using an ultrafiltration unit, and the ultrafiltration retentate is an enzymatic bone powder aqueous solution;

[0111] S6. The ultrafiltration permeate produced in S5 is treated in the manner of S4.1 to produce a nanofiltration permeate and a nanofiltration retentate;

[0112] S7. The nanofiltration permeate produced in steps S4 and S6 is transferred to a reverse osmosis transfer tank for reverse osmosis. The reverse osmosis retentate is concentrated to a solid content of 25% and then transferred to a nanofiltration retentate transfer tank. The mixture is then vacuum concentrated to a solid content of more than 40% and then spray dried to obtain amino acid powder.

[0113] S8. The nanofiltration retentate produced by the treatment in S4.1 is first vacuum concentrated (temperature 50-60 degrees Celsius) to a solid content of 50%, and then transferred to a spray drying transfer tank. The spray drying inlet air temperature is 170-190°C, and the outlet air temperature is 80-90°C to obtain bovine collagen peptides.

[0114] S9. The ultrafiltration retentate produced by S5 is concentrated to a water content of 50-70%, and then spray-dried (spray drying inlet air temperature 170-190°C, outlet air 80-90°C) to obtain enzymatic bone powder;

[0115] Or the ultrafiltration retentate is concentrated to a solid content of 10%, then reverse osmosis water is added to 1 times the amount and concentrated again to a solid content of 10%, and then 60-65% ethanol is used for alcohol precipitation, the precipitate is dehydrated with alcohol, granulated, dried at 75°C, crushed, and sieved through an 80-mesh sieve to obtain enzymatic bone powder.

[0116] The products obtained in Examples 1 and 2 were tested

[0117] Collagen peptides are tested according to the national standard GB31645;

[0118] Sodium hyaluronate: refer to QB / T4576-2013 for testing;

[0119] More than 80% of the number average molecular weight of sodium chondroitin sulfate is less than 10,000. The detection method is in accordance with the appendix of GB31645;

[0120] Sodium chondroitin sulfate and protein impurities USP (United States Pharmacopoeia): Sodium chondroitin sulfate: content not less than 90%, protein not higher than 6%; the content of sodium chondroitin sulfate, protein and sodium hyaluronate is tested on enzymatic bone powder.

[0121] The results are as follows:

[0122]

[0123] It can be seen that the yield of sodium chondroitin sulfate produced by the process of the present invention based on animal cartilage is as high as 5-9%, which is significantly higher than that reported in the prior art; the yield of collagen peptide is about 14%.

Claims

1. A production process of enzymatic bone powder rich in low molecular weight sodium chondroitin sulfate, characterized in that: The following steps are involved: S1. Pretreatment: The animal cartilage is steamed, and then the floating oil is removed, and then the animal cartilage is crushed in an extraction tank; the extraction tank is provided with a stirring paddle and a baffle with a shearing function at the bottom, the tank body is in a downward cone shape, and the bottom cover has a water inlet pipe and a valve at the bottom; the animal cartilage is crushed into fine particles by shearing with a stirring paddle with a shearing function and squeezing between bone particles during the stirring process, and the overall liquid is in a slurry state; S2. Enzymatic hydrolysis: Cool the liquid to 50-55 degrees, adjust the pH to 7-8, and then add protease for 0.5-2 hours. S3. Degreasing by adsorption: the enzymatic solution is released from the upper part of the extraction tank to the degreasing tank for degreasing by adsorption; S4. Ceramic membrane filtration: The deoiled liquid is collected in the first extract transfer tank and then pumped into the ceramic membrane filter for filtration; S5. Circulate the retentate filtered by the ceramic membrane: reflux the retentate filtered by the ceramic membrane into the second extract transfer tank, that is, the feed liquid in the second extract transfer tank is introduced into the extract tank through the bottom water inlet pipe of the extract tank to perform the enzymatic hydrolysis in step S2; S3 the enzymatic hydrolyzate passes through the bone granular layer in the extract tank and is released from the top to flow into the deoiling tank; S4 the feed liquid after adsorption and deoiling is collected in the first extract transfer tank, and then pumped into the ceramic membrane filter for filtration, and the generated retentate filtered by the ceramic membrane continues to circulate and reflux to the second extract transfer tank; The circulation process is continued until there are no obvious cartilage particles in the liquid material in the second extraction liquid transfer tank, and the circulation is terminated when the liquid material is clear; During the circulation process, reverse osmosis water is added to the second extract transfer tank to make up for the water lost by the ceramic membrane filtration; S6. After the circulation treatment is completed, the liquid in the second extraction liquid transfer tank is pumped back to the extraction tank through the reflux pump, and the stirring speed of the extraction tank is increased. After 20-80 minutes, the liquid in the extraction tank is pumped into the self-unloading flat plate centrifuge, and the filtrate is collected through the extraction transfer tank, and then added to the cation exchange resin tank through the carbon removal filter. After soaking for 2-5 hours, it is pumped into the ultrafiltration liquid transfer tank for subsequent ultrafiltration using an ultrafiltration unit. The ultrafiltration retentate is an enzymatic bone powder aqueous solution.

2. The production process according to claim 1, further comprising preparing enzymatic bone powder based on the ultrafiltration retentate produced by S6 by one or both of the following methods: (1) concentrating the ultrafiltration retentate to a water content of 50-70%, and then spray-drying to obtain enzymatic bone powder; (2) After the ultrafiltration retentate is concentrated to 10%, reverse osmosis water is added to concentrate it to 10% again, and then alcohol precipitation, dehydration, granulation, drying, crushing and sieving are performed to obtain enzymatic bone powder.

3. A production process based on animal cartilage to produce collagen peptides, sodium chondroitin sulfate and amino acid powder, characterized in that: The following steps are involved: S1. Pretreatment: The animal cartilage is steamed, and then the floating oil is removed, and then the animal cartilage is crushed in an extraction tank; the extraction tank is provided with a stirring paddle and a baffle with a shearing function at the bottom, the tank body is in a downward cone shape, and the bottom cover has a water inlet pipe and a valve at the bottom; the animal cartilage is crushed into fine particles by shearing with a stirring paddle with a shearing function and squeezing between bone particles during the stirring process, and the overall liquid is in a slurry state; S2. Enzymatic hydrolysis: cool the liquid to 50-55 degrees, adjust the pH to 7-8, and then add protease for 0.5-2 hours; S3. Degreasing by adsorption: the enzymatic solution is discharged from the upper part of the extraction tank to the degreasing tank for degreasing by adsorption; S4. Ceramic membrane filtration: The deoiled liquid is collected in the first extract transfer tank and then pumped into the ceramic membrane filter for filtration; S4.1 The permeate filtered by the ceramic membrane is pumped into a nanofiltration concentrator with a 300 Dalton nanofiltration membrane for nanofiltration. After the concentration reaches 15%, 1 volume of reverse osmosis water is added for further concentration. The nanofiltration retentate is temporarily stored in a nanofiltration retentate transfer tank, and then vacuum concentrated and dried to obtain bovine collagen peptides. The nanofiltration permeate is stored in a storage tank. S4.2 The retentate filtered by the ceramic membrane is circulated: the retentate filtered by the ceramic membrane is refluxed and introduced into the second extract transfer tank, that is, the feed liquid in the second extract transfer tank is introduced into the extraction tank through the bottom water inlet pipe of the extraction tank to perform the enzymatic hydrolysis in step S2; S3 The enzymatic hydrolyzate passes through the bone granular layer in the extraction tank and is released from the top to flow into the deoiling tank; S4 The feed liquid after adsorption and deoiling is collected in the first extract transfer tank, and then pumped into the ceramic membrane filter for filtration. The ceramic membrane filtration retentate produced continues to circulate back to the second extraction liquid transfer tank; the ceramic membrane filtration permeate produced in the circulation treatment is treated in the manner of S4.1 to produce nanofiltration permeate; The circulation process is continued until there are no obvious cartilage particles in the liquid material in the second extraction liquid transfer tank, and the circulation is terminated when the liquid material is clear; During the circulation process, reverse osmosis water is added to the second extract transfer tank to make up for the water lost by the ceramic membrane filtration; S5. The second extract transfer tank liquid is pumped back to the extraction tank through the reflux pump, the stirring speed of the extraction tank is increased, and the liquid in the extraction tank is pumped into the self-unloading flat centrifuge after 20-80 minutes, and the filtrate is collected by the extraction transfer tank, and then added to the cation exchange resin tank through the carbon removal filter, and then soaked for 2-5 hours and pumped into the ultrafiltrate transfer tank for subsequent ultrafiltration using an ultrafiltration unit, and the ultrafiltration retentate is an enzymatic bone meal aqueous solution; S6. The ultrafiltration permeate produced in S5 is treated in the same manner as in S4.1 to produce a nanofiltration permeate; S7. The nanofiltration permeate produced in steps S4 and S6 is transferred to a reverse osmosis transfer tank for reverse osmosis. The reverse osmosis retentate is concentrated to 25% and then vacuum concentrated to a solid content of more than 40%, and then spray-dried to obtain amino acid powder.

4. The production process according to any one of claims 1 to 3, wherein: The protease used in S2 is a mixed enzyme of 10000U / g flavor protease and 200000U / g alkaline protease in a mass ratio of 2:1, and the addition ratio is 0.5-0.9% of the mass of the feed liquid; the filter membrane parameter of the ceramic membrane filter is 3000 Dalton.

5. According to the production process of claim 3, the drying temperature of the vacuum concentration and drying of the nanofiltration retentate in step 4.1 is controlled to be 170-190 degrees Celsius for the inlet air temperature and 80-90 degrees Celsius for the outlet air temperature.

6. The production process according to claim 3, further comprising drying and crushing the filter residue produced by centrifugation in step S5 to obtain high calcium powder.

7. The production process according to claim 3, further comprising collecting the oil removed in steps S1 and S3.

8. The production process according to any one of claims 1 to 7, characterized in that: The animal cartilage includes one or more of bovine cartilage, chicken cartilage, fish cartilage, and pig cartilage.

9. The production process according to any one of claims 1 to 7, characterized in that: The extraction tank has a cylindrical tank body, and the lower part of the tank body is conical; a motor is arranged on the top of the tank body, and a stirring shaft controlled by the motor is located at the axis center position in the tank body and extends to the bottom of the cone; a stirring paddle with a shearing function is arranged at the lower part of the stirring shaft, and stirring paddles are arranged in the middle and upper parts; The bottom cover of the tank body is provided with a bottom water inlet pipe, a valve, and an air inlet pipe; the top of the tank body is provided with a discharge port for discharging the enzymatic hydrolysis liquid into the oil removal tank. Preferably, the top of the tank body is also provided with a feeding hole, an alkali pipe, and an acid pipe; A lower temperature detection tube, a middle temperature detection tube and an upper temperature detection tube are sequentially arranged on the side wall of the tank from bottom to top; A sampling and analysis tube 16 and a disinfectant tube 17 are also provided on the side wall of the tank body 1 .

10. An equipment set for producing enzymatic bone powder rich in sodium chondroitin sulfate, characterized in that: It includes any of the above extraction tanks, an oil removal tank, a first extraction liquid transfer tank, a ceramic membrane filter and a second extraction liquid transfer tank connected by a pipeline closed loop, and is used for cyclic processing of raw materials; The extraction tank is used to stir, crush and hydrolyze animal cartilage with enzymes; the extraction tank has a cylindrical tank body, and the lower part of the tank body is conical; a motor is arranged on the top of the tank body, and a stirring shaft controlled by the motor is located at the axis position of the tank body and extends to the bottom of the cone; a stirring paddle with a shearing function is arranged at the lower part of the stirring shaft, and stirring paddles are arranged in the middle and upper parts; The bottom cover of the tank body is provided with a bottom water inlet pipe, a valve, and an air inlet pipe; the top of the tank body is provided with a discharge port for discharging the enzymatic hydrolysis liquid into the degreasing tank; The liquid inlet of the de-oiling tank is connected to the discharge port provided at the upper part of the extraction tank through a pipeline, so that the enzymatic hydrolysis liquid in the extraction tank can be discharged into the de-oiling tank for de-oiling; The first extraction liquid transfer tank is used to collect the deoiled liquid from the deoiling tank. The ceramic membrane filter is used to filter the liquid from the first extraction liquid transfer tank. The second extraction liquid transfer tank is used to collect the filtered intercepted liquid produced by the ceramic membrane filter, which is connected to the bottom water inlet pipe 6 at the bottom of the extraction tank through a pipeline, so that the filtered intercepted liquid is returned to the extraction tank through the bottom water inlet pipe 6 by a reflux pump, thereby realizing the circulation treatment of the feed liquid; In a preferred embodiment, the equipment combination further comprises an ultrafiltration transfer tank and an ultrafiltration device, wherein the ultrafiltration transfer tank is used to collect the slurry that has completed the circulation treatment in the extraction tank, and the ultrafiltration device is used to ultrafilter the slurry that has completed the circulation treatment, and the intercepted liquid produced by the ultrafiltration is post-treated to obtain the enzymatic bone powder; Preferably, a feeding hole, an alkali pipe and an acid pipe are also provided on the top of the tank body; A lower temperature detection tube, a middle temperature detection tube and an upper temperature detection tube are arranged on the side wall of the tank body from bottom to top in sequence; a sampling and analysis tube and a disinfectant liquid tube are also arranged on the side wall of the tank body 1.

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