Hydrolyzed silk protein preparation method and integrated reaction device
By integrating the silk fibroin preparation unit into an integrated reaction device, and employing saponin degumming, ultrasonic-assisted salt dissolution, and compound enzymatic hydrolysis, the problem of low production efficiency in existing technologies has been solved, realizing the efficient preparation and industrial production of silk fibroin.
Patent Information
- Application Number
- CN202511014631.3
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-07-23
- Publication Date
- 2025-10-28
AI Technical Summary
Existing silk protein preparation equipment has limited functionality, requiring each process to be completed in different devices, resulting in low production efficiency and making it difficult to achieve large-scale industrial production. Furthermore, processes such as degumming, dissolving, desalting, and enzymatic hydrolysis suffer from low efficiency and poor results.
An integrated reaction device was designed, which integrates the washing, drying, crushing and separation and hydrolysis of silk protein preparation units on the workbench. It adopts saponin degumming, ultrasonic-assisted salt dissolution, nanofiltration desalination and compound enzymatic hydrolysis. The auxiliary components realize automatic material transfer and simplify the operation process.
It improves production efficiency, reduces equipment footprint, and lowers the intensity of manual operation, enabling safe, simple, and economical large-scale industrial production. Furthermore, the prepared silk protein has a small molecular weight, high activity, and excellent quality.
Smart Images
Figure CN120843263A_ABST
Abstract
Description
Technical Field
[0001] This invention relates to the field of silk fibroin processing equipment technology, specifically to a method for preparing hydrolyzed silk fibroin and an integrated reaction device. Background Technology
[0002] Silk fibroin has excellent biocompatibility and biodegradability, and has broad application prospects in the fields of medicine, cosmetics, and food.
[0003] However, existing silk fibroin preparation devices are often single-function, requiring each process to be completed in different devices, resulting in low production efficiency and making it difficult to achieve large-scale industrial production. At the same time, there are many problems in the degumming, dissolving, desalting, and enzymatic hydrolysis processes during the preparation of silk fibroin, such as poor degumming effect, low dissolving efficiency, long desalting time, and low enzymatic hydrolysis efficiency, which affect the quality and yield of silk fibroin. To address these issues, this invention proposes a method for preparing hydrolyzed silk fibroin, an integrated reaction device, and a preparation method to solve the above problems. Summary of the Invention
[0004] The purpose of this invention is to provide a method for preparing hydrolyzed silk protein and an integrated reaction apparatus to solve the problems mentioned in the background art.
[0005] To achieve the above objectives, the present invention provides the following technical solution: a method for preparing hydrolyzed silk protein and an integrated reaction apparatus, including a workbench;
[0006] The workbench is equipped with a cleaning unit, a drying unit, a pulverizing and separating unit, and an integrated unit for preparing hydrolyzed silk protein.
[0007] The cleaning unit includes a cleaning tank, an ultrasonic generator is provided on the inner wall of the cleaning tank, a drain cavity is provided at the bottom of the cleaning tank, a water pump is provided in the drain cavity, the water pump is connected to the collection cylinder with a drain pipe, and a perforated plate is provided above the drain cavity.
[0008] Preferably, an auxiliary component is connected through the cleaning unit, drying unit, and crushing and separating unit. The auxiliary component includes a U-shaped support plate and a first cylinder, with the first cylinder connected to the U-shaped support plate.
[0009] Preferably, the drying unit includes an oven, the inner cavity of which can accommodate the laterally movable U-shaped support plate, the temperature of which is controlled at 40-60℃, and a hot air circulating heating method is adopted.
[0010] Preferably, the crushing and separating unit includes a slitting blade holder and a second cylinder. The slitting blade holder extends laterally with three sets of assembly areas. Cutting blades are installed below the three sets of assembly areas, and the three sets of cutting blades are equally spaced at 3cm intervals.
[0011] Preferably, the integrated unit for preparing hydrolyzed silk protein specifically includes a degumming unit, a salt dissolution unit, a desalting unit, an enzymatic hydrolysis unit, a decolorization unit, and a low-temperature concentration unit.
[0012] Preferably, the degumming unit includes a reaction pressure vessel, the upper end of which is provided with a first marking block, and the reaction pressure vessel is capable of maintaining a constant pressure of 15 psi and a temperature of 100°C.
[0013] Preferably, the salt dissolving unit includes an ultrasonic-assisted heating centrifugal dissolving device, which consists of an ultrasonic generator, a heating device, and a centrifugal device, and the upper cover of the ultrasonic-assisted heating centrifugal dissolving device is provided with a second marking block.
[0014] Preferably, the desalination unit uses a nanofiltration membrane cartridge, and the top cover of the nanofiltration membrane cartridge is provided with a third marking block.
[0015] Preferably, the enzymatic hydrolysis unit includes a water bath oscillation heating inactivation integrated device, which is composed of a water bath oscillation device and a heating inactivation device, and the upper end of the water bath oscillation heating inactivation integrated device is provided with a fourth marking block.
[0016] A method for preparing hydrolyzed silk protein, comprising the following specific steps:
[0017] Step 1: Cleaning and drying. Clean the cut silkworm cocoons with clean water to remove impurities, and dry them at a low temperature of 40-60℃.
[0018] Step 2: Crushing and separating, crushing the dried, shaved silkworm cocoons into 2-4cm cocoon fragments;
[0019] Step 3: Degumming. Add 0.024 g / cc saponin aqueous solution to the silk sample and soak it in the reaction pressure vessel. Remove the insoluble degummed silk fibroin, clean it, and dry it.
[0020] Step 4: Salt dissolution. The dried degummed silk fibroin is dissolved by ultrasonication in the ternary system solution. Finally, it is cooled to room temperature and centrifuged to obtain a crude silk fibroin mixture.
[0021] Step 5: Desalination. The crude silk protein mixture is desalted using nanofiltration to obtain a solution free of calcium. 2+ Mg 2+ Cl - Silk protein solution;
[0022] Step 6: Enzymatic hydrolysis. Add 0.65% compound enzyme to the silk fibroin solution, and hydrolyze by shaking in a water bath at 35-37℃. Then boil the hydrolysate to inactivate it, centrifuge and filter to obtain the supernatant.
[0023] Step 7: Decolorization. Use activated carbon to decolorize the supernatant to obtain a silk protein solution.
[0024] Step 8: Low-temperature concentration. The silk protein solution is concentrated at 55°C until the solid content is 25%-40%, resulting in a concentrated silk protein solution.
[0025] Compared with the prior art, the beneficial effects of the present invention are:
[0026] (1) By integrating the washing, drying, crushing and separation and hydrolyzed silk protein preparation units on the workbench, the equipment footprint is reduced. The materials are automatically transferred between the units through auxiliary components, eliminating the repeated transfer of hydrolyzed silk between different processing containers and reducing the intensity of manual operation. At the same time, by integrating the entire process of hydrolyzed silk protein preparation, the production process is safe, simple, convenient and economical, and easy to industrialize and mass-produce.
[0027] (2) By using saponin to degumme silkworm cocoons, no other chemical reagents need to be added. As a natural biosurfactant, the surface activity of saponin molecules can effectively remove sericin protein in silk fibroin and keep the surface of silk fibroin protein smooth and the surface roughness unchanged, thereby reducing the toxicity of silk fibroin protein extract.
[0028] (3) By cooperating with the first cylinder through the U-shaped support plate of the auxiliary component, the automatic transfer of materials between the cleaning, drying and crushing separation units is realized, avoiding pollution and loss caused by manual material transfer, and ensuring the cleanliness and integrity of material processing.
[0029] (4) The hydrolyzed silk protein prepared by this device has a small molecular weight range, around 1000-3000. The small molecular weight results in high activity.
[0030] (5) The production process of the present invention is safe, simple, convenient, and economical, and is easy to scale up for industrial production. Saponin is used to degumm the silkworm cocoons without the addition of other chemical reagents. Saponin is an excellent degumming agent, and the surface activity of saponin molecules can effectively remove sericin from silk fibroin. Due to the addition of natural biosurfactant saponin, the surface of silk fibroin remains smooth, the surface roughness remains unchanged, and the toxicity of the silk fibroin extract is reduced. Ultrasonic assistance during the salt dissolution process can accelerate the dissolution efficiency of silk fibroin. The use of nanofiltration for desalination solves the problems of long desalination time, low desalination efficiency, and unsuitability for industrial production of conventional dialysis bags. Serai peptide enzyme and Alcalase protease are used to enzymatically hydrolyze the silk fibroin solution. Serai peptide enzyme is produced by fermentation of Serratia marcescens isolated from the intestine of silkworm pupae and has a strong protein hydrolysis ability. Numerous studies have shown that Alcalase is the enzyme with the highest yield in hydrolyzing silk protein. Therefore, the combination of serrepeptidase and Alcalase solves the current problems of low hydrolysis efficiency and low activity of silk protein. The hydrolyzed silk protein produced by this method has a relatively small molecular weight range, around 1000-3000, which is small in molecular weight but has high activity. Attached Figure Description
[0031] Figure 1 This is a schematic diagram of the overall structure of the present invention.
[0032] Figure 2 This is a schematic diagram of the structure from another perspective of the present invention.
[0033] Figure 3 This is a schematic diagram of the cleaning unit structure of the present invention.
[0034] Figure 4 This is a schematic diagram of the installation structure of the cleaning unit and auxiliary components of the present invention.
[0035] Figure 5 This is a schematic diagram of the crushing and separating unit structure of the present invention.
[0036] Figure 6 This is a schematic diagram of the integrated unit structure for preparing hydrolyzed silk protein according to the present invention.
[0037] In the diagram: 1. Workbench; 2. Cleaning unit; 21. Cleaning tank; 211. Ultrasonic generator; 22. Drainage chamber; 23. Partition plate; 24. Collection cylinder; 3. Drying unit; 4. Crushing and separation unit; 41. Slitting blade holder; 42. Second cylinder; 5. Auxiliary components; 51. U-shaped support plate; 52. First cylinder; 6. Hydrolyzed silk protein preparation integrated unit; 61. Degumming unit; 62. Salt dissolution unit; 63. Desalination unit; 64. Enzymatic hydrolysis unit; 65. Decolorization unit; 66. Low-temperature concentration unit. Detailed Implementation
[0038] 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 of ordinary skill in the art without creative effort are within the scope of protection of the present invention.
[0039] Please see Figures 1 to 6 The present invention provides a technical solution: an integrated reaction device for hydrolyzing silk protein, including a workbench 1, on which a cleaning unit 2, a drying unit 3, a crushing and separation unit 4 and a hydrolyzed silk protein preparation integrated unit 6 are provided. The cleaning unit 2 includes a cleaning tank 21, an ultrasonic generator 211 is provided on the inner wall of the cleaning tank 21, a drainage chamber 22 is provided at the bottom of the cleaning tank 21, a water pump is provided in the drainage chamber 22, the water pump is connected to the collection cylinder 24 with a drainage pipe, and a perforated plate 23 is provided above the drainage chamber 22.
[0040] By integrating the washing, drying, crushing and separation and hydrolyzed silk protein preparation units on the workbench 1, the equipment footprint is reduced. The auxiliary components 5 enable automatic material transfer between the units, eliminating the need for repeated transfer of hydrolyzed silk between different processing containers and reducing the intensity of manual operation. At the same time, by integrating the entire process of hydrolyzed silk protein preparation, the production process is safe, simple, convenient, economical, and easy to scale up for industrial production.
[0041] Please see Figures 1-4 It is understood that an auxiliary component 5 is connected through the cleaning unit 2, the drying unit 3, and the crushing and separating unit 4. The auxiliary component 5 includes a U-shaped support plate 51 and a first cylinder 52, and the first cylinder 52 is connected to the U-shaped support plate 51.
[0042] Please see Figure 1-2 It is understood that the drying unit 3 includes an oven, the inner cavity of which can accommodate a laterally movable U-shaped support plate 51, the temperature of the oven is controlled at 40-60℃, and a hot air circulating heating method is adopted.
[0043] Please see Figure 1 , Figure 5 It is understood that the crushing and separating unit 4 includes a slitting blade holder 41 and a second cylinder 42. The slitting blade holder 41 extends laterally with three sets of assembly areas. Cutting blades are installed below the three sets of assembly areas, and the three sets of cutting blades are evenly spaced at 3cm intervals.
[0044] Please see Figure 1-6 It is understood that the integrated unit 6 for preparing hydrolyzed silk protein specifically includes a degumming unit 61, a salt dissolution unit 62, a desalting unit 63, an enzymatic hydrolysis unit 64, a decolorization unit 65, and a low-temperature concentration unit 66.
[0045] Please see Figure 2 , Figure 6 It is understood that the degumming unit 61 includes a reaction pressure vessel. In the degumming unit 61 of the present invention, the reaction pressure vessel capable of maintaining a constant pressure of 15 psi and a temperature of 100°C can be selected, such as the GSH series high-pressure reactor produced by Weihai Huanyu Chemical Machinery Co., Ltd. This model of reactor is made of high-quality stainless steel and has good pressure resistance and temperature resistance. At the same time, the reactor body is equipped with a stirring device, which can fully mix 10 grams of silk sample with 200 ml of 0.024 g / cc saponin aqueous solution during the degumming process, ensuring the uniformity and efficiency of degumming and improving the degumming effect.
[0046] Please see Figure 2 It is understood that the salt dissolving unit 62 includes an ultrasonic-assisted heating centrifugal dissolving device, which consists of an ultrasonic generator, a heating device, and a centrifugal device. The upper cover of the ultrasonic-assisted heating centrifugal dissolving device is provided with a second marking block.
[0047] Please see Figure 2 It is understood that the desalination unit 63 uses a nanofiltration membrane cartridge, and a third marking block is provided at the top cover of the nanofiltration membrane cartridge.
[0048] Please see Figure 2 Understandably, the enzymatic digestion unit 64 includes a water bath oscillation heating inactivation integrated device, which consists of a water bath oscillation device and a heating inactivation device. A fourth marking block is provided on the upper end of the water bath oscillation heating inactivation integrated device.
[0049] Please see Figure 2 , Figure 6 It is understood that the decolorization unit 65 and the low-temperature concentration unit 66 respectively include a stirring tank and a low-temperature concentration tank, wherein the upper cover of the stirring tank and the low-temperature concentration tank are respectively provided with a fifth marking block and a sixth marking block.
[0050] Working principle:
[0051] First, the cut silkworm cocoons are placed onto the U-shaped support plate 51. At this time, the U-shaped support plate 51 is in the cleaning tank 21 of the cleaning unit 2. The cut silkworm cocoons are cleaned by adding clean water to the cleaning tank 21. After the impurities are removed by the clean water, the water pump in the drain chamber 22 at the lower end of the cleaning tank 21 is started to pump the water from the drain chamber 22 and the cleaning tank 21 into the collection cylinder 24. Then, the first cylinder 52 is started, which moves the U-shaped support plate 51 to the drying unit 3. It is worth noting that both ends of the U-shaped support plate 51 are tightly filled with the side walls of the cleaning tank 21 to ensure a seal. When the cut silkworm cocoons are transferred to the drying unit 3, the drying unit is started to dry them at a temperature of 40-60℃. The dried cocoons then enter the crushing and separating unit 4. At this point, the first cylinder 52 is activated, moving the U-shaped support plate 51 and the cut silkworm cocoons into the crushing and separating unit 4. Then, the second cylinder 42 is activated, moving the cutting blade holder 41 downwards. Since the three sets of cutting blades are evenly spaced at 3cm intervals, the dried cut silkworm cocoons can be crushed into 3cm cocoon fragments. The chopped cocoon fragments are removed from the crushing and separating unit 4 and poured into the hydrolyzed silk protein preparation integrated unit 6 for further processing.
[0052] First, the silkworm cocoon fragments enter the degumming unit 61 and are soaked in 200 ml of 0.024 g / cc saponin aqueous solution at a constant pressure of 15 psi and 100 °C for 24 minutes. The degummed insoluble silk fibroin is then removed and dried by the dryer in the reaction pressure vessel.
[0053] Firstly, the dried degummed silk fibroin enters the salt dissolution unit 62, is mixed with the ternary system solution in proportion, and is ultrasonically dissolved for 40 minutes at 50℃ and 30kw. After cooling, it is centrifuged to obtain a crude silk fibroin mixture.
[0054] Secondly, the crude silk protein mixture enters the desalination unit 63, where it is desalinated by nanofiltration to obtain a silk protein solution.
[0055] Third, the silk fibroin solution enters the enzymatic hydrolysis unit 64, 0.65% of the complex enzyme is added, and the solution is hydrolyzed in a water bath at 35-37℃ for 3 hours with shaking. Then it is boiled for 35 minutes to inactivate the enzyme, and the supernatant is collected by centrifugation and filtration.
[0056] Fourth, the supernatant enters the decolorization unit 65, where it is mixed with 3% activated carbon for decolorization, resulting in a clear and transparent silk protein solution.
[0057] Finally, the silk fibroin solution enters the low-temperature concentration unit 66 and is concentrated at 55°C to a solid content of 25%-40%, yielding a concentrated silk fibroin solution.
[0058] It is worth noting that the reaction vessels of the degumming unit 61, salt dissolution unit 62, desalting unit 63, enzymatic hydrolysis unit 64, decolorization unit 65, and low-temperature concentration unit 66 are marked with serial numbers to prevent errors in the preparation process.
[0059] Although embodiments of the invention have been shown and described, it will be understood by those skilled in the art that various changes, modifications, substitutions and alterations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the appended claims and their equivalents.
Claims
1. An integrated reaction apparatus for hydrolyzing silk protein, comprising a workbench (1), characterized in that: The workbench (1) is equipped with a cleaning unit (2), a drying unit (3), a crushing and separation unit (4), and an integrated unit for preparing hydrolyzed silk protein (6); The cleaning unit (2) includes a cleaning tank (21), an ultrasonic generator (211) is provided on the inner wall of the cleaning tank (21), a drain cavity (22) is provided at the bottom of the cleaning tank (21), a water pump is provided in the drain cavity (22), the water pump is connected to the collection cylinder (24) with the drain pipe, and a partition plate (23) is provided above the drain cavity (22).
2. The integrated reaction apparatus for hydrolyzing silk fibroin according to claim 1, characterized in that: An auxiliary component (5) is connected through the cleaning unit (2), drying unit (3), and crushing and separating unit (4). The auxiliary component (5) includes a U-shaped support plate (51) and a first cylinder (52), which is connected to the U-shaped support plate (51).
3. The integrated reaction apparatus for hydrolyzing silk fibroin according to claim 2, characterized in that: The drying unit (3) includes an oven, the inner cavity of which can accommodate the laterally movable U-shaped support plate (51), the temperature of which is controlled at 40-60℃, and a hot air blower is used for circulating heating.
4. The integrated reaction apparatus for hydrolyzing silk fibroin according to claim 2, characterized in that: The crushing and separating unit (4) includes a slitting blade holder (41) and a second cylinder (42). The slitting blade holder (41) extends laterally with three sets of assembly areas. Cutting blades are installed below the three sets of assembly areas, and the three sets of cutting blades are evenly spaced at 3-5cm intervals.
5. The integrated reaction apparatus for hydrolyzing silk fibroin according to claim 1, characterized in that: The hydrolyzed silk protein preparation integrated unit (6) specifically includes a degumming unit (61), a salt dissolution unit (62), a desalting unit (63), an enzymatic hydrolysis unit (64), a decolorization unit (65), and a low-temperature concentration unit (66).
6. The integrated reaction apparatus for hydrolyzing silk fibroin according to claim 5, characterized in that: The degumming unit (61) includes a reaction pressure vessel, the upper end of which is provided with a first marking block, and the reaction pressure vessel is capable of maintaining a constant pressure of 15 psi and a temperature of 100°C.
7. The integrated reaction apparatus for hydrolyzing silk fibroin according to claim 5, characterized in that: The salt dissolving unit (62) includes an ultrasonic-assisted heating centrifugal dissolving device, which consists of an ultrasonic generator, a heating device and a centrifugal device. The upper cover of the ultrasonic-assisted heating centrifugal dissolving device is provided with a second marking block.
8. The integrated reaction apparatus for hydrolyzing silk fibroin according to claim 5, characterized in that: The desalination unit (63) adopts a nanofiltration membrane cartridge, and a third marking block is provided on the top cover of the nanofiltration membrane cartridge.
9. The integrated reaction apparatus for hydrolyzing silk fibroin according to claim 5, characterized in that: The enzymatic hydrolysis unit (64) includes a water bath oscillation heating inactivation integrated device, which is composed of a water bath oscillation device and a heating inactivation device. The upper end of the water bath oscillation heating inactivation integrated device is provided with a fourth marking block.
10. A method for preparing hydrolyzed silk protein, characterized in that, The specific steps are as follows: Step 1: Cleaning and drying. Clean the cut silkworm cocoons with clean water to remove impurities, and dry them at a low temperature of 40-60℃. Step 2: Crushing and separating, crushing the dried, shaved silkworm cocoons into 2-4cm cocoon fragments; Step 3: Degumming. Add 0.024 g / cc saponin aqueous solution to the silk sample and soak it in the reaction pressure vessel. Remove the insoluble degummed silk fibroin, clean it, and dry it. Step 4: Salt dissolution. The dried degummed silk fibroin is dissolved by ultrasonication in the ternary system solution. Finally, it is cooled to room temperature and centrifuged to obtain a crude silk fibroin mixture. Step 5: Desalination. The crude silk protein mixture is desalted using nanofiltration to obtain a solution free of calcium. 2+ Mg 2+ Cl - Silk protein solution; Step 6: Enzymatic hydrolysis. Add 0.65% compound enzyme to the silk fibroin solution, and hydrolyze by shaking in a water bath at 35-37℃. Then boil the hydrolysate to inactivate it, centrifuge and filter to obtain the supernatant. Step 7: Decolorization. Use activated carbon to decolorize the supernatant to obtain a silk protein solution. Step 8: Low-temperature concentration. The silk protein solution is concentrated at 55°C until the solid content is 25%-40%, resulting in a concentrated silk protein solution.