Method for separating and recovering active protein from whey

Abstract

The invention provides a method for separating and recovering active protein from whey, which comprises the following steps: adding a certain amount of chitosan into the whey, adjusting the pH value of the solution, and centrifugally precipitating to obtain a precipitate B and a clear liquid C; adding a sodium chloride solution with a certain concentration into the precipitate B, adjusting the pH value of the precipitate B, redissolving the precipitate B to obtain a supernatant E rich in beta-lactoglobulin, and drying the supernatant E to obtain a high-purity beta-lactoglobulin finished product; and synchronously concentrating and desalting the alpha-lactalbumin in the clear liquid C by using a separation device formed by coupling membrane filtration and electrodialysis, and drying to obtain a high-purity alpha-lactalbumin finished product. According to the method for separating and recovering the active protein from the whey, provided by the invention, high-purity beta-lactoglobulin and alpha-lactalbumin finished products can be simply, conveniently, quickly and efficiently obtained from the whey waste liquid, the investment and operation cost of a separation system for the active protein in the whey is reduced, the treatment time is saved, and industrial amplification is facilitated.

Description

Technical Field

[0001] The invention belongs to the technical field of active protein separation and recovery, and in particular relates to a method for separating and recovering high-purity alpha-lactalbumin and beta-lactoglobulin from whey. Background Art

[0002] Whey wastewater discharged during cheese or yogurt production is the main source of pollution in the dairy industry, and whey wastewater contains a high content of whey protein with rich nutritional value and biological activity. Whey protein is a mixture of various functional proteins, the main components of which are α-lactalbumin, β-lactoglobulin, bovine serum albumin, lactoferrin and immunoglobulin, of which α-lactalbumin accounts for 20-25% of the total protein content, and β-lactoglobulin accounts for 30-55% of the total protein content. α-Lactalbumin has the functional properties of anti-hypertension, antioxidant activity, anti-cancer and anti-obesity, and can be widely used in the research and development of functional health products and infant formula milk powder. β-Lactoglobulin also has the effects of anti-hypertension, anti-cancer and appetite enhancement, but β-lactoglobulin is the main factor inducing milk allergy. Therefore, how to effectively separate α-lactalbumin and β-lactoglobulin in whey to obtain high-purity α-lactalbumin and β-lactoglobulin respectively can not only effectively reduce the environmental pollution of whey wastewater, but also has unique biological and medical value.

[0003] At present, the commonly used methods for separating and recovering α-lactalbumin and β-lactoglobulin from whey include precipitation method by heating or changing pH to selectively cause protein precipitation, membrane technology using multi-stage ultrafiltration for separation, and ion exchange chromatography. The precipitation method requires a large amount of acid, alkali and salt, the protein is severely denatured, and the post-processing process is complicated. Although the ion exchange chromatography can separate α-lactalbumin and β-lactoglobulin with higher purity, the yield and application scale are difficult to further expand and the energy consumption is high. Although the membrane technology has low energy consumption and is easy to industrialize, it is difficult to effectively separate α-lactalbumin and β-lactoglobulin with similar molecular weight. The Chinese patent with publication number CN 118271424 B discloses a method for separating and preparing fresh milk grade high-purity α-lactalbumin. A higher purity α-lactalbumin solution is obtained by combining microfiltration and anion exchange chromatography purification. The method has a simple preparation process and is easy to industrialize, but it fails to achieve effective separation of α-lactalbumin and β-lactoglobulin. The Chinese patent with publication number CN 104256053 B discloses a method for preparing α-lactalbumin powder and β-lactoglobulin powder, which mainly adopts pH adjustment and multi-stage ultrafiltration technology to obtain two active protein powders respectively. Although this method reduces membrane pollution and improves membrane separation efficiency, the protein purity is low, and the protein is easy to change during pH adjustment. At present, the methods for separating α-lactalbumin and β-lactoglobulin from whey have the problems of high energy consumption, low protein purity, and inability to effectively separate α-lactalbumin and β-lactoglobulin to varying degrees. Therefore, it is urgent to develop a simple, efficient, and easy to industrially scale up method for separating and recovering active proteins in whey. Summary of the invention

[0004] The purpose of the present invention is to provide a method for separating and recovering α-lactalbumin and β-lactoglobulin from whey that is simple, efficient, and easy to scale up industrially. The present invention first separates and recovers β-lactalbumin from whey by adding a certain amount of chitosan to whey, utilizing the interaction between chitosan and β-lactoglobulin in whey to form a precipitate, and re-dissolving the precipitate formed by chitosan and β-lactoglobulin at a certain salt solution concentration and pH; then a porous filter membrane with a certain pore size is set in a conventional electrodialysis membrane stack to construct a new type of device formed by intrinsic coupling of membrane filtration and electrodialysis for further separation and recovery of α-lactalbumin in whey. The method achieves high-purity α-lactalbumin and β-lactoglobulin from whey by a two-step method combining chitosan extraction and electromembrane process separation. Compared with the existing α-lactalbumin and β-lactoglobulin separation technology, the process is simple, easy to scale up industrially, and can achieve the co-recovery of α-lactalbumin and β-lactoglobulin.

[0005] The objective of the present invention is achieved through the following technical solutions:

[0006] A method for separating and recovering active protein from whey, firstly, chitosan is added to whey, the concentration of chitosan and the pH of the solution are adjusted to enhance the binding force between chitosan and β-lactoglobulin in whey, and then after standing and centrifugation, a precipitate and a clarified liquid formed by chitosan and β-lactoglobulin are obtained. According to the principle that the interaction mode and size between chitosan and β-lactoglobulin are closely related to the salinity and pH of the solution, a NaCl solution of a certain concentration is added to the precipitate formed by chitosan and β-lactoglobulin, and its pH is adjusted to re-dissolve the chitosan and β-lactoglobulin precipitate, thereby obtaining high-purity β-lactoglobulin. The separation and extraction of β-lactoglobulin eliminates interference with the subsequent separation and recovery of α-lactalbumin, and the clarified liquid rich in α-lactalbumin obtained by the above centrifugation is pumped into a separation and recovery device formed by the internal coupling of membrane filtration and electrodialysis to simultaneously concentrate and desalt the α-lactalbumin in the whey. The membrane stack of the separation device used includes positive and negative electrode chambers on both sides, the inner sides of which are two electrode chamber protection chambers, and an α-lactalbumin recovery chamber and a feed and liquid chamber composed of anion exchange membrane, recovery chamber partition, porous filter membrane, feed and liquid chamber partition, which are arranged alternately. The clarified liquid rich in α-lactalbumin obtained after separation of β-lactoglobulin by chitosan enters the feed and liquid chamber. Under the drive of the DC electric field and the selective separation of the porous filter membrane and the anion exchange membrane, the negatively charged α-lactalbumin in the feed and liquid chamber will migrate across the porous filter membrane to the anode direction and enter the recovery chamber, while the bovine serum albumin, lactoferrin, immunoglobulin, etc. in the feed and liquid chamber are retained in the feed and liquid chamber due to the pore size screening effect of the porous membrane, and then carried out of the feed and liquid chamber with the liquid flow; the small molecular salt positive and negative ions in the α-lactalbumin recovery chamber migrate into the adjacent feed and liquid chamber under the action of the electric field, thereby realizing the synchronous separation and desalination of α-lactalbumin. The solution in the α-lactalbumin recovery chamber is circulated in a closed loop, wherein the concentration of α-lactalbumin increases continuously, and finally an α-lactalbumin concentrate is obtained; after further drying of the concentrate, a high-purity α-lactalbumin finished product can be obtained.

[0007] The above method for separating and recovering active protein from whey is characterized by being carried out according to the following steps:

[0008] (1) adding a certain amount of chitosan solution to whey A, adjusting the pH of the solution to a specific value, standing for 5-20 minutes, and then centrifuging for 10 minutes to obtain a precipitate B and a clarified liquid C;

[0009] (2) adding a certain concentration of NaCl solution to the precipitate B obtained in step (1) and adjusting the pH thereof to 9.0-12.0 to dissolve the precipitate to obtain a solution D in which β-lactoglobulin is redissolved;

[0010] (3) centrifuging the solution D obtained in step (2) at a speed of 3000-6000 r / min for 5-20 min to obtain a supernatant E rich in β-lactoglobulin;

[0011] (4) pumping the clarified liquid C obtained in step (1) as a raw liquid into an α-lactalbumin separation and recovery device formed by the internal coupling of membrane filtration and electrodialysis, and simultaneously concentrating and desalting the α-lactalbumin in the clarified liquid C to obtain a concentrated liquid F rich in α-lactalbumin;

[0012] (5) The supernatant E rich in β-lactoglobulin obtained in step (3) and the concentrated solution F rich in α-lactalbumin obtained in step (4) are subjected to spray drying, low-temperature vacuum drying or freeze drying to obtain high-purity β-lactoglobulin and α-lactalbumin finished products.

[0013] Preferably, the chitosan described in step (1) is chitosan of low or medium viscosity.

[0014] Preferably, the amount of chitosan added in step (1) is 1.0-5.0 mg / mL.

[0015] Preferably, after adding the chitosan solution in step (1), the pH of the solution is adjusted to 4.0-8.0, preferably to 6.5.

[0016] The membrane stack of the above-mentioned α-lactalbumin separation and recovery device is composed of a number of repeatedly arranged basic working units, wherein each basic working unit includes an α-lactalbumin recovery chamber and a feed liquid chamber composed of an anion exchange membrane and a porous filter membrane, and the separation side of the porous filter membrane faces the cathode.

[0017] Preferably, the molecular weight cut-off of the porous filter membrane used in the α-lactalbumin separation and recovery device in step (4) is 20,000-50,000.

[0018] Preferably, the material of the porous filter membrane used in the α-lactalbumin separation and recovery device described in step (4) can be organic polymer materials such as cellulose, polysulfone, polyvinylidene fluoride, etc., or inorganic materials such as ceramics, metals, molecular sieves, etc.

[0019] Preferably, the thickness of the liquid chamber and the egg albumin recovery chamber of the α-lactalbumin separation and recovery device in step (4) are both 0.5-5.0 mm.

[0020] Preferably, the positive and negative electrode chambers of the α-lactalbumin separation and recovery device described in step (4) are both provided with electrode chamber protection chambers.

[0021] Preferably, the electrode liquid and electrode protection liquid of the α-lactalbumin separation and recovery device described in step (4) are both separately prepared 5.0-20.0 g / L Na 2 SO 4 The solution first enters the anode chamber and the anode protection chamber from the lower part of the anode chamber and the anode protection chamber respectively, is discharged at the upper part, and then enters the cathode chamber and the cathode protection chamber from the lower part of the cathode chamber and the cathode protection chamber respectively through the external pipeline of the membrane stack, and is finally discharged at the upper part of the cathode chamber and the cathode protection chamber, forming independent electrode liquid and protection liquid pipelines.

[0022] In the present invention, a chitosan solution obtained by dissolving in 0.1 mol / L hydrochloric acid is added to whey A, and the addition amount is controlled to be 1.0-5.0 mg / mL. Then, the pH of the solution is adjusted to 4.0-8.0, so that chitosan is positively charged and β-lactoglobulin is negatively charged at the pH, and an insoluble precipitate is formed by the electrostatic attraction between the two. After standing for 5-20 minutes, the mixture is centrifuged for 10 minutes to obtain a chitosan and β-lactoglobulin composite precipitate B and a clear solution C. The charge of chitosan changes with the pH of the solution. When the pH of the solution is alkaline, chitosan is negatively charged, the interaction between β-lactoglobulin and chitosan is converted from electrostatic attraction to electrostatic repulsion, the chitosan and β-lactoglobulin complex is dissociated, chitosan is precipitated, and β-lactoglobulin is redissolved in the solution. Furthermore, a certain concentration of NaCl solution is added to the chitosan and β-lactoglobulin composite precipitate B to adjust its pH to 9.0-12.0, so that the precipitate can be dissolved, and the β-lactoglobulin solution D can be obtained by re-dissolving. Furthermore, the supernatant E rich in β-lactoglobulin is obtained by centrifugation at a speed of 3000-6000 r / min for 5-20 min.

[0023] In the present invention, the clear liquid C obtained after adding chitosan precipitation treatment is pumped into the α-lactalbumin separation and recovery device formed by the intrinsic coupling of membrane filtration and electrodialysis as the raw liquid for treatment to obtain the α-lactalbumin concentrate. Further, the membrane stack of the α-lactalbumin separation and recovery device in whey is composed of a plurality of basic working units consisting of an anion exchange membrane and a porous filter membrane, which are repeatedly arranged. The porous filter membrane adopts an ultrafiltration membrane with a molecular weight of 20,000-50,000, allowing α-lactalbumin to pass freely while being able to effectively intercept the transmembrane migration of macromolecular proteins such as lactoferrin and immunoglobulin coexisting therewith. The clear liquid C is pumped into the feed liquid chamber of the α-lactalbumin separation and recovery device as the raw liquid. Under the dual effects of the aperture screening of the porous filter membrane and the electric field, the negatively charged egg α-lactalbumin in the clear liquid C will cross the porous filter membrane and enter the α-lactalbumin recovery chamber on the left, and the α-lactalbumin concentrate F is obtained with the multiple closed-loop circulation of the solution in the α-lactalbumin recovery chamber.

[0024] The arrangement of the anode protection chamber in the present invention can, on the one hand, prevent H, one of the anode reaction products, from + ions migrate and diffuse to the first α-lactalbumin recovery chamber adjacent to it, thereby maintaining the stable existence of α-lactalbumin in the recovery chamber; on the other hand, it can also prevent the Cl - Ions migrate to the anode chamber and produce harmful gas Cl 2 The cathode protection chamber can prevent the cathode reaction product OH - The migration of ions to the last feed liquid chamber close to the cathode side maintains the stability of the pH value of the feed liquid chamber, thereby ensuring the migration of α-lactalbumin in the feed liquid from the feed liquid chamber to the recovery chamber under the action of the electric field.

[0025] The electrode solution and electrode protection solution in the present invention are both separately prepared 5.0-20.0 g / L Na 2 SO 4 Solution to maintain good conductivity of the cathode, anode and protection chambers. The electrode liquid and electrode protection liquid enter from the anode side, are discharged from the cathode side, and enter the external electrode liquid circulation tank and electrode protection liquid circulation tank. After removing the gas generated by the electrode reaction, they can be recycled. During normal operation, due to the electrode reaction, the liquid flow in the anode chamber is acidic, and the liquid flow in the cathode chamber is alkaline. The electrode liquid is introduced from the anode chamber to the cathode chamber, and the accumulation of electrode reaction products can be eliminated due to neutralization to eliminate the impact on the adjacent feed liquid chamber and α-lactalbumin recovery chamber.

[0026] In the present invention, by using feed and liquid chambers and α-lactalbumin recovery chamber baffles of different area specifications, and increasing or decreasing the number of basic working units composed of α-lactalbumin recovery chambers and feed and liquid chambers in the device, whey of different scales can be processed, which is convenient for industrial scale-up.

[0027] The method for separating and recovering active protein from whey according to the present invention has the following effects:

[0028] (1) Chitosan solution in whey A, by adjusting the pH of the solution to control the interaction mode and size of chitosan and β-lactoglobulin, forming a composite precipitate B of β-lactoglobulin and chitosan, and re-dissolving the precipitate to obtain a solution rich in β-lactoglobulin, and after drying, obtaining a β-lactoglobulin finished product. This method can not only achieve efficient recovery of β-lactoglobulin in whey, but also obtain a β-lactoglobulin finished product with high purity.

[0029] (2) The effective integration of chitosan extraction technology for β-lactoglobulin and α-lactalbumin separation technology formed by the intrinsic coupling of membrane filtration and electrodialysis can respectively obtain high-purity α-lactalbumin and β-lactoglobulin products from whey, thereby achieving efficient recovery of both.

[0030] (3) The scale of the recovery process of α-lactalbumin in whey can be adjusted by adopting partition plates with different area specifications for the recovery chamber and the feed solution chamber, and increasing or decreasing the number of basic working units composed of the recovery chamber and the feed solution chamber, which is convenient for industrial scale-up.

[0031] (4) Compared with the existing technologies of chromatographic separation, multi-stage ultrafiltration separation process and their integration, the process flow is simpler. The α-lactalbumin and β-lactoglobulin concentrates obtained by the method for separating and recovering active proteins from whey provided in the present invention can be directly dried without further desalination or resin adsorption. The recovered α-lactalbumin and β-lactoglobulin have high activity and purity, the system investment is saved, the process is simple, the time consumption is short and the operation stability is high. Description of the Drawings

[0032] Figure 1 It is a schematic diagram of a method for separating and recovering active proteins from whey provided by the present invention;

[0033] Figure 2 It is a schematic diagram of the internal structure of the α-lactalbumin separation and recovery device provided by the present invention;

[0034] Figure 3 It is a specific process flow diagram of an embodiment device provided by the present invention.

[0037] In the above figures:

[0038] 1 - porous filter membrane; 2 - anion exchange membrane; 3 - feed solution chamber; 4 - α-lactalbumin recovery chamber; 5 - feed solution at the inlet of the feed solution chamber; 6 - solution at the inlet of the α-lactalbumin recovery chamber; 7 - anode; 8 - cathode; 9 - cation exchange membrane; 10 - anode chamber; 11 - anode protection chamber; 12 - cathode protection chamber; 13 - cathode chamber; 14 - basic working unit; 15 - water inlet of the electrode chamber; 16 - water outlet of the electrode chamber; 17 - solution at the outlet of the α-lactalbumin recovery chamber; 18 - solution at the outlet of the feed solution chamber; 19 - water inlet of the protection chamber; 20 - water outlet of the protection chamber; 21 - power supply; 22 - α-lactalbumin separation and recovery device; 23 - feed solution circulation tank; 24 - α-lactalbumin recovery solution circulation tank; 25 - electrode solution circulation tank; 26 - electrode protection solution circulation tank; 27 - feed solution circulation pump; 28 - α-lactalbumin recovery solution circulation pump; 29 - electrode solution circulation pump; 30 - electrode protection solution circulation pump; 31 - stop valve; 32 - rotameter; 33 - on-line pH meter. Detailed Embodiments

[0039] To facilitate the understanding of the technical solution of the present invention by those skilled in the art, the technical solution of the present invention will be further specifically described below in conjunction with the accompanying drawings of the specification and embodiments.

[0040] like Figure 1 As shown, the present invention is a method for separating and recovering active proteins from whey, comprising a two-step method for separating and recovering α-lactalbumin and β-lactoglobulin from whey, namely, firstly adding a certain amount of chitosan to the whey, and then adjusting the pH of the solution to form a composite precipitate of chitosan and β-lactoglobulin and a clarified liquid rich in α-lactalbumin; adding a certain concentration of NaCl solution to the composite precipitate of chitosan and β-lactoglobulin, and adjusting the pH of the solution to be alkaline, so that the β-lactoglobulin in the precipitate is redissolved, and centrifugal precipitation can obtain a supernatant rich in β-lactalbumin; and the clarified liquid rich in α-lactalbumin enters an α-lactalbumin separation and recovery device for separation and enrichment to obtain an α-lactalbumin concentrated liquid; the supernatant rich in β-lactalbumin and the α-lactalbumin concentrated liquid are dried to obtain high-purity β-lactoglobulin and α-lactalbumin finished products. The specific operation steps are:

[0041] (1) adding 1.0-5.0 mg / mL chitosan solution to whey A, adjusting the pH of the solution to 4.0-8.0, standing for 5-20 min, and then centrifuging for 10 min to obtain precipitate B and clarified liquid C;

[0042] (2) adding a certain concentration of NaCl solution to the precipitate B obtained in step (1) and adjusting the pH thereof to 9.0-12.0 to dissolve the precipitate to obtain a solution D in which β-lactoglobulin is redissolved;

[0043] (3) centrifuging the solution D obtained in step (2) at a speed of 3000-6000 r / min for 5-20 min to obtain a supernatant E rich in β-lactoglobulin;

[0044] (4) pumping the clarified liquid C obtained in step (1) as a raw liquid into an α-lactalbumin separation and recovery device formed by the internal coupling of membrane filtration and electrodialysis, and simultaneously concentrating and desalting the α-lactalbumin in the clarified liquid C to obtain a concentrated liquid F rich in α-lactalbumin;

[0045] (5) The supernatant E rich in β-lactoglobulin obtained in step (3) and the concentrated solution F rich in α-lactalbumin obtained in step (4) are subjected to spray drying, low-temperature vacuum drying or freeze drying to obtain high-purity β-lactoglobulin and α-lactalbumin finished products.

[0046] according to Figure 2The basic working unit 14 of the membrane stack of a separation and recovery device 22 for α-lactalbumin in whey, which is composed of a porous filter membrane 1, an anion exchange membrane 2 and a cation exchange membrane 9, includes a feed liquid chamber 3 and an α-lactalbumin recovery chamber 4. Under the action of the external electric field provided by the power supply 21, the negatively charged α-lactalbumin in the feed liquid chamber 3 migrates to the α-lactalbumin recovery chamber 4 adjacent to the left side through the porous filter membrane 1; the α-lactalbumin is retained in the α-lactalbumin recovery chamber 4 due to the control of the solution pH and the pore size screening and selective permeability of the anion exchange membrane. The α-lactalbumin retained in the α-lactalbumin recovery chamber 4 will be carried out of the membrane stack with the liquid flow, and the synchronous desalination and high-multiple concentration of the α-lactalbumin will be achieved through multiple cycles. The porous filter membrane 1 arranged in the liquid chamber 3 has a molecular weight cutoff of 20,000-50,000, so that the negatively charged α-lactalbumin molecules in the liquid chamber 3 can be driven by an external electric field and smoothly migrate through the porous filter membrane 1 into the adjacent α-lactalbumin recovery chamber 4 on the left, thereby achieving efficient separation and enrichment and recovery of α-lactalbumin in whey.

[0047] according to Figure 3 The provided process flow is that the clear liquor C obtained by adding chitosan precipitation treatment as the feed liquid chamber inlet solution 5 enters the α-lactalbumin separation and recovery device 22 formed by the intrinsic coupling of membrane filtration and electrodialysis through the feed liquid circulation tank 23 and the feed liquid circulation pump 27, flows through the feed liquid chamber 3 from bottom to top, and then returns to the feed liquid circulation tank 23 to perform closed-loop circulation, so that the α-lactalbumin in the whey continuously migrates into the α-lactalbumin recovery liquid. The α-lactalbumin recovery chamber inlet solution 6 is pumped into the membrane stack by the α-lactalbumin recovery liquid circulation pump 28, flows through the α-lactalbumin recovery chamber 4 from bottom to top, and then returns to the α-lactalbumin recovery liquid circulation tank 24 to perform closed-loop circulation, so as to continuously absorb the α-lactalbumin from the clear liquor C, so as to achieve the high multiple concentration of α-lactalbumin. The electrode chamber inlet water 15 enters the anode chamber 10 and the cathode chamber 13 from bottom to top from the electrode liquid circulation tank 25 through the electrode liquid circulation pump 29. The electrode chamber outlet water 16 returns to the electrode liquid circulation tank 25 to discharge gas and then circulates into the anode chamber 10 and the cathode chamber 13 as the electrode chamber inlet water 15. The protection chamber inlet water 19 enters the anode protection chamber 11 and the cathode protection chamber 12 from bottom to top from the electrode protection liquid circulation tank 26 through the electrode liquid circulation pump 30. The protection chamber outlet water 20 returns to the electrode protection liquid circulation tank 26 as the electrode protection chamber inlet water 19 and circulates into the anode protection chamber 11 and the cathode protection chamber 12.

[0048] Example

[0049] In this embodiment, 3.0 g of whey protein powder is added to a NaCl solution with a concentration of 2000 mg / L, and after sufficient stirring and dissolving, a simulated whey with a total protein content of 3000 mg / L is obtained, to which a low-viscosity chitosan solution dissolved by 0.1 mol / L HCl is added, and the addition amount of chitosan is 2.0 mg / mL. The pH is adjusted to 6.5, and the mixture is sufficiently stirred for 15 min. After standing for 10 min, the mixture is centrifuged at 6000 r / min in a high-speed centrifuge for 10 min to obtain a composite precipitate of chitosan and β-lactoglobulin and a clarified liquid.

[0050] After the composite precipitate of chitosan and β-lactoglobulin was separated, 4.0 mol / L NaCl was added to the precipitate, and the pH of the solution was adjusted to 11.0 with a NaOH solution. The β-lactoglobulin was re-dissolved by stirring, and the supernatant rich in β-lactoglobulin was obtained at a speed of 6000 r / min for 10 min. The β-lactoglobulin product was obtained after freeze-drying, and was detected as β-lactoglobulin by high performance liquid chromatography.

[0051] The clarified liquid obtained after the simulated whey was treated with chitosan precipitation entered the α-lactalbumin separation and recovery device formed by the intrinsic coupling of membrane filtration and electrodialysis to further separate and concentrate the α-lactalbumin therein, and the obtained α-lactalbumin recovery liquid was vacuum freeze-dried, and the obtained α-lactalbumin finished product was detected as α-lactalbumin by high performance liquid chromatography.

[0052] The α-lactalbumin separation device is a one-stage structure, containing a basic working unit, which is used to separate α-lactalbumin from the clarified liquid after chitosan precipitation treatment. The specifications of the feed liquid chamber, α-lactalbumin recovery chamber, cathode and anode chamber protection chamber, and cathode and anode chamber partitions are all 1500mm×750mm×9mm. The ion exchange membrane used is a heterogeneous ion exchange membrane produced by Zhejiang Qianqiu Environmental Protection Water Treatment Co., Ltd. The porous filter membrane used is a cellulose acetate ultrafiltration membrane with a molecular weight cutoff of 50,000, provided by Ande Membrane Separation Technology Engineering Co., Ltd.

[0053] The liquid circulation tank 23 contains the clarified liquid obtained by adding chitosan to simulate whey to adjust pH, stirring and standing, and centrifuging. The initial solution in the α-lactalbumin recovery liquid circulation tank 24 is a 2000 mg / L NaCl solution; the electrode liquid and the protective liquid are 20.0 g / L Na 2 SO 4The pH values ​​of the solution, feed solution and α-lactalbumin recovery solution are monitored by an online acidometer 33, and the content and purity of α-lactalbumin are determined by high performance liquid chromatography. The flow rates of the feed solution, α-lactalbumin recovery solution, electrode solution and electrode protection solution are 30.0, 30.0, 100.0 and 100.0 L / min respectively, and the membrane stack working voltage is 6V. After 5.0 hours of stable operation, the recovery rate of α-lactalbumin in whey can reach 26.1%, the purity can reach 97%, and the recovery rate of β-lactoglobulin can reach 82.4%, and the purity can reach 94.6%. The embodiment shows that by utilizing the active protein separation and recovery method in whey provided by the present invention, high-purity α-lactalbumin and β-lactoglobulin can be obtained from whey in a simple and effective manner. This new α-lactalbumin and β-lactoglobulin separation and recovery technology can greatly improve the yield of active proteins in whey, reduce the investment and operating costs of the protein separation system, and has important application value in the fields of separation and purification of active proteins.

Claims

1. A method for separating and recovering active protein from whey, characterized in that Follow these steps: (1) adding a certain amount of chitosan solution to whey A, adjusting the pH of the solution to a specific value, standing for 5-20 minutes, and then centrifuging for 10 minutes to obtain a precipitate B and a clarified liquid C; (2) adding a certain concentration of NaCl solution to the precipitate B obtained in step (1) and adjusting the pH thereof to 9.0-12.0 to dissolve the precipitate to obtain a solution D in which β-lactoglobulin is redissolved; (3) centrifuging the solution D obtained in step (2) at a speed of 3000-6000 r / min for 5-20 min to obtain a supernatant E rich in β-lactoglobulin; (4) pumping the clarified liquid C obtained in step (1) as a raw liquid into an α-lactalbumin separation and recovery device formed by the internal coupling of membrane filtration and electrodialysis, and simultaneously concentrating and desalting the α-lactalbumin in the clarified liquid C to obtain a concentrated liquid F rich in α-lactalbumin; (5) The supernatant E rich in β-lactoglobulin obtained in step (3) and the concentrated solution F rich in α-lactalbumin obtained in step (4) are subjected to spray drying, low-temperature vacuum drying or freeze drying to obtain high-purity β-lactoglobulin and α-lactalbumin finished products. The α-lactalbumin separation and recovery device described in step (4) comprises four parts: a membrane stack, an electrode device, a clamping support device and a clamping device, wherein the membrane stack is composed of a plurality of repeatedly arranged basic working units, characterized in that each basic working unit comprises a feed liquid chamber composed of an anion exchange membrane and a porous filter membrane, and an α-lactalbumin recovery chamber, wherein the separation side of the porous filter membrane faces the cathode, the molecular weight cutoff of the porous filter membrane used is 20000-50000, and a polar chamber protection chamber is provided on the inner side of the positive and negative electrode chambers.

2. A method for separating and recovering active protein from whey according to claim 1, characterized in that The chitosan described in step (1) is chitosan of low or medium viscosity.

3. A method for separating and recovering active protein from whey according to claim 1, characterized in that The amount of chitosan added in step (1) is 1.0-5.0 mg / mL.

4. A method for separating and recovering active protein from whey according to claim 1, characterized in that After adding the chitosan solution in step (1), the pH of the solution is adjusted to 4.0-8.0, preferably to 6.

5.

5. A method for separating and recovering active protein from whey according to claim 1, characterized in that The material of the porous filter membrane used in the α-lactalbumin separation and recovery device described in step (4) can be organic polymer materials such as cellulose, polysulfone, polyvinylidene fluoride, etc., or inorganic materials such as ceramics, metals, molecular sieves, etc.

6. A method for separating and recovering active protein from whey according to claim 1, characterized in that The thickness of the liquid chamber and the α-lactalbumin recovery chamber of the α-lactalbumin separation and recovery device in step (4) is 0.5-5.0 mm.

7. A method for separating and recovering active protein from whey according to claim 1, characterized in that The electrode liquid and electrode protection liquid of the α-lactalbumin separation and recovery device described in step (4) are both separately prepared 5.0-20.0 g / L Na2SO4 solutions, and first enter the anode chamber and the anode protection chamber from the lower parts of the anode chamber and the anode protection chamber respectively, are led out at the upper part, and then enter the cathode chamber and the cathode protection chamber from the lower parts of the cathode chamber and the cathode protection chamber respectively through the external pipeline of the membrane stack, and are finally discharged at the upper parts of the cathode chamber and the cathode protection chamber, forming independent electrode liquid and protection liquid pipelines.

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