CN112679593B - Method for recovering sarcoplasmic proteins in minced fillet rinsing liquid by combining acid treatment with chitosan flocculation and application - Google Patents
Method for recovering sarcoplasmic proteins in minced fillet rinsing liquid by combining acid treatment with chitosan flocculation and application Download PDFInfo
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Abstract
The invention relates to a method for recovering sarcoplasmic proteins in minced fillet rinsing liquid by combining acid treatment with chitosan flocculation and application thereof, wherein fish is placed in distilled water for rinsing and centrifugation, and the supernatant is the minced fillet rinsing liquid; adding chitosan solution into the minced fillet rinsing liquid, firstly adjusting the pH to 2.0-4.0, then adjusting the pH to 6.0-9.0 by NaOH solution, standing, centrifuging, and precipitating to obtain the co-flocculate of the sarcoplasmic protein and the chitosan. Compared with the traditional chitosan flocculation method, the recovery rate of the sarcoplasmic protein by chitosan can be improved by more than one time through acid treatment, the recovered sarcoplasmic protein and chitosan co-flocculate does not need to be separated, the good spinnability is presented, the sarcoplasmic protein co-flocculate can be directly applied to electrostatic spinning to manufacture a sarcoplasmic protein nano fibrous membrane, and the electro-spinning fibrous membrane has strong surface hydrophobicity, mechanical property and thermal stability. The invention can save energy and reduce consumption, reduce the treatment cost and pressure of the minced fillet rinsing wastewater, and solve the problems of recovery and high-value utilization of byproducts in the industrialized production of the minced fillet.
Description
Technical Field
The invention belongs to the technical field of food, and relates to a method for recovering sarcoplasmic proteins in minced fillet rinsing liquid by combining acid treatment and chitosan flocculation and application thereof.
Background
The minced fillet product is very high in nutritional value and tender mouthfeel due to the fact that the minced fillet product is rich in high-quality protein and amino acid, is widely popular worldwide, and the demand of the minced fillet product is continuously increased all the year round. In 2019, the total amount of processed aquatic products in China is 2171.41 ten thousand tons, and the yield of the minced fillet and the minced fillet products is about 139.40 ten thousand tons. The silver carp has high yield, low price and fresh and tender meat, and is a main freshwater fish for producing the minced fillet. However, in the production process of minced fillet, a large amount of water is needed to rinse the fish for multiple times to remove the residual impurities such as blood, lipid, water-soluble protein and sarcolemma, on one hand, the water-soluble sarcoplasmic protein accounting for 20-40% of the fish protein is lost in the rinsing process, which causes serious waste of resources; on the other hand, a large amount of rinsing wastewater rich in protein is generated and needs to be treated, otherwise, eutrophication of the water body is caused. Therefore, if the sarcoplasmic proteins in the minced fillet rinsing liquid can be fully recovered, the utilization rate of the sarcoplasmic proteins and the added value of fish products can be improved, organic pollutants in the rinsing wastewater can be reduced, the treatment cost is reduced, and good economic value and environmental benefit are generated.
Chitosan is a natural straight-chain high-molecular polymer, the main chain of which contains abundant amino groups, can be protonated in an acidic medium, shows the characteristic of a typical cationic polyelectrolyte and endows the chitosan with good flocculation performance. In addition, chitosan has good biocompatibility and degradability, so that the chitosan is widely used in wastewater treatment. However, the chitosan has poor solubility in neutral or alkaline aqueous solution, which prevents the optimal use of the chitosan, so that the recovery rate of the sarcoplasmic proteins in the minced fillet rinsing liquid is low, namely about 40 percent by using the chitosan alone. Therefore, how to improve the flocculation effect of chitosan on sarcoplasmic proteins and improve the protein recovery rate becomes a problem to be solved urgently.
Disclosure of Invention
In order to solve the technical problems, the invention provides a method for recovering sarcoplasmic proteins in minced fillet rinsing liquid by combining acid treatment and chitosan flocculation and application thereof.
The scheme of the invention is as follows:
a method for recovering sarcoplasmic proteins in minced fillet rinsing liquid by acid treatment and chitosan flocculation, which comprises the following steps:
s1: preparing a minced fillet rinsing liquid: adding distilled water into fish meat for rinsing, centrifuging, and taking supernatant as minced fillet rinsing liquid;
s2: adding chitosan solution into the minced fillet rinsing liquid, and adjusting the pH value to 6.0-9.0;
s3: standing, centrifuging, and precipitating to obtain the co-flocculate of sarcoplasmic protein and chitosan.
Preferably, in the step S1, the solid-to-liquid ratio of the fish meat to the distilled water is 1 (1.0-3.5) (g/mL), and the centrifugation conditions are as follows: 8000-11000g,4 deg.C, 12-18min.
Preferably, in the step S2, the deacetylation degree of the chitosan is 85-98%, and the addition amount of the chitosan in the minced fillet rinsing liquid is 150-400mg/L.
Preferably, in the step S2, the pH is adjusted to 2.0-4.0, and then adjusted to 6.0-9.0 by NaOH solution.
Further preferably, in the step S2, the pH is adjusted to 3.0, and then adjusted to 7.0 by using NaOH solution.
Still more preferably, in the step S2, the concentration of the NaOH solution is 1.0-6.0mol/L.
Preferably, in the step S3, the step of placing the mixture into a water bath kettle for standing is performed at a temperature of 10-35 ℃, and after the standing time is 30-300min, the centrifugation conditions are as follows: 3500-4500r/min,8-15min.
The sarcoplasmic proteins recovered by the method are used for manufacturing the sarcoplasmic proteins electrostatic spinning nanofiber membrane, and the application comprises the following steps:
1) Freeze-drying sarcoplasmic protein and chitosan, dissolving in hexafluoroisopropanol, magnetically stirring,
obtaining a spinning solution;
2) Carrying out electrostatic spinning on the spinning solution, and receiving the fiber membrane by rolling aluminum foil paper;
3) And drying the fiber membrane to remove residual organic solvent to obtain the sarcoplasmic protein electrostatic spinning nanofiber membrane.
Preferably, the electrostatic spinning conditions of step 2) are as follows: electrostatic spinning is carried out at the temperature of 13-27 ℃, the humidity of 30-65 percent, under the conditions of using electrostatic voltage of 15-22KV, the spinning speed of 0.4-1.8mL/h and the distance between a spinning nozzle and a receiving plate of 10-18 cm.
Preferably, the drying conditions in step 3) are as follows: naturally drying at room temperature or drying in a vacuum drying oven at 35-60 deg.C.
The invention has the beneficial effects that:
1. compared with the traditional chitosan flocculation method, the acid treatment can improve the recovery rate of the chitosan to the sarcoplasmic proteins by more than one time, the action mechanism of the invention is derived from the protonation of the chitosan in an acid solution with positive charge, the chitosan has good solubility, and can be fully dissolved with the sarcoplasmic proteins; when the pH value is increased, protein molecules are gradually dissociated into an electric negative state, and are easily and rapidly subjected to electrostatic attraction with the positively charged chitosan to be flocculated and settled. The sarcoplasmic protein and chitosan co-flocculate recovered by the invention does not need to be separated, has good spinnability, can be directly applied to electrostatic spinning to prepare the sarcoplasmic protein nano fiber membrane, and the electrospun fiber membrane has strong surface hydrophobicity, mechanical property and thermal stability. The method for recovering the sarcoplasmic proteins by flocculation saves energy, reduces consumption, reduces the treatment cost and pressure of the minced fillet rinsing wastewater, and solves the problems of recovery and high-value utilization of byproducts in the industrial production of the minced fillet.
2. The acid treatment can improve the effect of flocculating the protein by the chitosan and also can well improve the removal of the chemical oxygen demand in the minced fillet rinsing liquid.
3. The acid treatment can obviously improve the recovery efficiency of the sarcoplasmic protein, and greatly reduces the energy consumption compared with the direct freeze drying method. Compared with the sarcoplasmic protein recovered by a single chitosan flocculation method, the sarcoplasmic protein flocculated by acid treatment and chitosan has good spinnability.
Drawings
FIG. 1 recovery of sarcoplasmic proteins by single chitosan flocculation;
FIG. 2 recovery of sarcoplasmic proteins by acid treatment in combination with chitosan flocculation;
FIG. 3 influence of pH on protein recovery;
FIG. 4 effect of chitosan dosage on protein recovery;
FIG. 5 effect of temperature on protein recovery;
FIG. 6 effect of time on protein recovery;
FIG. 7 is a scanning electron micrograph of different concentrations of flocculated sarcoplasmic protein electrospun membranes;
FIG. 8 contact angles of flocculated and directly freeze-dried sarcoplasmic protein fibrous membranes;
FIG. 9 mechanical properties of flocculated and direct freeze-dried sarcoplasmic protein fibrous membranes;
FIG. 10 DSC curves of chitosan powder and sarcoplasmic protein fibrous membranes in different recovery modes.
Detailed Description
The invention is further illustrated by the following examples, but the scope of the invention as claimed is not limited to the scope of the examples.
Example 1
1 materials and reagents
The farmer market of Huazhong agriculture university of silver carp; chitosan, beijing Kulaibo science and technology Limited, is dissolved in 1% by mass of acetic acid to prepare a 1% by mass chitosan solution (1 wt% chitosan solution) when in use.
2 method of experiment
2.1 preparation of surimi rinsing liquid [21]
Knocking dizzy of fresh silver carp, removing head, tail, skin and internal organs, manually picking meat, mincing with a conditioner, and weighing fish meat. Rinsing fish meat with 1.
2.2 recovery of sarcoplasmic proteins by Single Chitosan flocculation
And (3) adding 40mL of minced fillet rinsing liquid into each centrifugal tube, and sequentially adding 1.2mL of 1wt% chitosan solution with deacetylation degrees of 85%, 90% and 95% into the test tube. The pH was subsequently adjusted to 7.0 with 3mol/L NaOH solution. Standing in 20 deg.C water bath for 90min, centrifuging at 4000r/min for 10min, and collecting supernatant to measure protein concentration.
2.3 recovery of sarcoplasmic proteins by acid treatment in combination with Chitosan flocculation
And (3) adding 40mL of minced fillet rinsing liquid into each centrifugal tube, and sequentially adding 1.2mL of 1wt% chitosan solution with deacetylation degrees of 85%, 90% and 95% into the test tube. The pH was adjusted to 3.0, followed by 3mol/L NaOH solution to 7.0. Standing in 20 deg.C water bath for 90min, centrifuging at 4000r/min for 10min, and collecting supernatant to measure protein concentration.
2.4 Single factor test
2.4.1 Effect of pH on protein recovery
And (3) taking 10 centrifugal tubes, adding 40mL of minced fillet rinsing liquid into each centrifugal tube, and adding 1.2mL of 1wt% chitosan solution into the test tubes respectively. The pH was adjusted to 3.0, followed by adjusting the pH with 3mol/L NaOH solution to 4.5, 5.0, 5.5, 6.0, 6.5, 7.0, 7.5, 8.0, 8.5, 9.0 in that order. Standing in a water bath kettle at 20 deg.C for 90min, centrifuging at 4000r/min for 10min, and collecting supernatant to measure protein concentration.
2.4.2 Effect of Chitosan dosage on protein recovery
Taking 6 centrifugal tubes, adding 40mL of minced fillet rinsing liquid into each centrifugal tube, and then sequentially adding 0.6, 0.8, 1.0, 1.2, 1.4 and 1.6mL of 1wt% chitosan solution (counting the dosage of chitosan in minced fillet rinsing water to be 150, 200, 250, 300, 350 and 400 mg/L). The pH was adjusted to 3.0, followed by 3mol/L NaOH solution to 7.0. Standing in water bath at 20 deg.C for 90min, centrifuging at 4000r/min for 10min, collecting supernatant, and measuring protein concentration
2.4.3 Effect of temperature on protein recovery
And (3) taking 7 centrifugal tubes, adding 40mL of minced fillet rinsing liquid into each centrifugal tube, and adding 1.2mL of 1wt% chitosan solution into the test tubes respectively. The pH was adjusted to 3.0, followed by sequential adjustment of the pH to 7.0 with 3mol/L NaOH solution. Standing in water bath at 10, 20, 30, 40, 50, 60, and 70 deg.C for 90min, centrifuging at 4000r/min for 10min, and collecting supernatant to determine protein concentration.
2.4.4 Effect of time on protein recovery
And (3) taking 7 centrifugal tubes, adding 40mL of minced fillet rinsing liquid into each centrifugal tube, and adding 1.2mL of 1wt% chitosan solution into the test tubes respectively. The pH was adjusted to 3.0, followed by 3mol/L NaOH solution to 7.0. Standing in a water bath kettle at 20 deg.C for 30, 60, 90, 120, 150, 180, and 210min, centrifuging at 4000r/min for 10min, and collecting supernatant to measure protein concentration.
2.5 response surface optimization test
On the basis of a single-factor test, box-Bechnken center combination optimization Design is carried out by using Design-expert software, the total recovery rate of final protein is used as a test index, and a three-factor three-level response surface test is carried out on 3 factors which possibly have synergistic effects, namely chitosan amount, pH value and temperature. Wherein the levels of-1, 0 and 1 of the three factors are respectively set as follows: the A factor has pH values of 6.5, 7.0 and 7.5; the temperature of the B factor is 10, 20 and 30 ℃; the dosage of the C-factor chitosan is 250, 300 and 350mg/L.
2.6 recovery of protein
Protein concentration was determined according to the Folin phenol method (standard curve regression equation: y =0.0023x +0.0751, R2= 0.9931) and protein recovery was calculated as follows:
in the formula: c 0 The concentration of protein in the original minced fillet rinsing liquid is mg/mL; c 1 Protein concentration in supernatant after flocculation, mg/mL.
3 results and analysis
3.1 recovery of sarcoplasmic proteins by Single Chitosan flocculation
The results of single chitosan flocculation to recover sarcoplasmic proteins are shown in FIG. 1. The recovery rates of the sarcoplasmic proteins recovered by the three deacetylation degrees of chitosan are as follows in sequence: the degree of deacetylation is greater than 95% and greater than 90% and greater than 85%. The increase of the deacetylation degree can lead to the increase of amino groups, so that the density of positive charges is increased after protonation, the charge neutralization effect is improved, and the chitosan has a better flocculation effect. Therefore, the subsequent experiments selected chitosan with a degree of deacetylation of 95%. The recovery rate of the sarcoplasmic proteins obtained by the single chitosan flocculation method with the deacetylation degree of 95 percent is lower and is only about 35 percent.
3.2 recovery of sarcoplasmic proteins by acid treatment in combination with Chitosan flocculation
The results of acid treatment combined with the chitosan flocculation method for recovering sarcoplasmic proteins are shown in FIG. 2. The acid treatment can obviously improve the flocculation effect of the chitosan as a single flocculant, and the recovery rate of the sarcoplasmic proteins reaches 81.54 percent. Compared with a single chitosan flocculation method, the protein recovery rate is improved by 46.02%. It can be seen that the acid treatment (pH adjusted to 3.0) plays a crucial role in the recovery of surimi rinse solution protein by chitosan flocculation, because the acid-treated chitosan is positively charged by protonation, has good water solubility, and is fully miscible with sarcoplasmic protein; then the pH value is increased, the protein molecules are gradually dissociated into an electric negative state, and the electric negative state and the positively charged chitosan are quickly subjected to electrostatic attraction to flocculate and are jointly settled. Meanwhile, isoelectric precipitation of the protein is also accompanied.
3.3 Single factor Experimental results and analysis
3.3.1 Effect of pH on protein recovery
The effect of pH on protein recovery is shown in figure 3. The pH value has a more obvious influence on the chitosan flocculation process. The recovery increases with increasing pH at pH 4.5 to 7.0, reaches a maximum at pH 7.0 and then gradually decreases. It is possible that at lower pH values, the protein species, this ampholyte, is positively charged in acidic solution, while the chitosan is positively charged by protonation of the chitosan's amino groups in acidic solution, the electrostatic repulsion effect of both conferring relative dynamic stability to the system, resulting in lower recovery; when the pH is more than the optimum pH of 7.0, the protein is solubilized in an alkaline environment, and the recovery rate is lowered.
3.3.2 Effect of Chitosan dosage on protein recovery
The effect of chitosan dosage on protein recovery is shown in figure 4. With the increase of the addition amount of chitosan, the protein recovery rate tended to increase and then decrease, reaching a maximum value when the chitosan amount was 300mg/L. It is possible that the reason is that as the chitosan content increases, chitosan and protein are able to undergo effective charge neutralization, bridging and electrostatic interaction to aggregate sedimentation, and the flocculation effect is optimal when the solution as a whole is electrically neutral. However, the amount of chitosan is too much, and the flocculation recovery rate of protein tends to be reduced.
3.3.3 Effect of temperature on recovery of sarcoplasmic proteins by Chitosan flocculation
The effect of temperature on protein recovery is shown in figure 5. When the temperature is below 30 ℃, the recovery rate does not change significantly with temperature, and there is a relatively high recovery rate at 20 ℃. When the temperature exceeds 30 ℃, the protein recovery rate tends to increase, and the possible reason is that the denaturation temperature of the sarcoplasmic proteins is about 40 ℃, the protein is thermally denatured due to high temperature, and the spatial structure is destroyed to destabilize and precipitate. Overall, it is not reasonable to trade a large increase in energy consumption for a small increase in recovery, so the optimum temperature for the flocculation process should be chosen to be 20 ℃.
3.3.4 Effect of time on recovery of sarcoplasmic proteins by Chitosan flocculation
The effect of time on protein recovery is shown in figure 6. With increasing flocculation time, protein recovery first tended to increase, reaching a maximum at 90min, after which recovery decreased. The possible reason is that chitosan is not sufficiently chelated with protein and flocculates to form a precipitate at the initial stage of flocculation; and the flocculation time is too long, the chitosan-protein aggregate is unstable and the depolymerization phenomenon occurs, so that the recovery rate is reduced.
3.4 response surface optimization experiment results and analysis
According to the result of the single-factor experiment, the flocculation time is set to 90min. The pH, the temperature and the chitosan content are respectively set as three investigation factors, the Box-Bechnken center combination optimization Design of Design-expert software is adopted, the total protein recovery rate is used as an investigation index for testing, and the result is shown in Table 1. Performing multiple quadratic regression fitting on the data in the table 1 to obtain a regression equation:
Y＝84.08-1.13A-0.12B+0.24C-0.17AB+1.08AC-0.082BC-0.072B 2 -0.39C 2 。
TABLE 1 Box-Bechnken test design and results
As can be seen from Table 2, the A and AC terms have a very significant effect on the experimental results by variance test and significance analysis (P)<0.01). Model Pr>F has a very significant effect on the experimental results (P)<0.01 The distortion term Pr>F value of 0.1064, and no significant effect on the experiment (P)>0.05 Showing that the model fitting degree is good and the variation in the response can be accurately simulated. Coefficient of determination R of model 2 Is 0.9201, illustrating this equation pairThe test fitting degree is good, and the error is small; properly modified R to increase the reliability of model prediction 2 The value of adj is 0.8401, which shows that the model can explain the change of 84.01% response value, and proves that the model can be used for fitting the relation between the independent variable and the response value of the experiment.
TABLE 2 analysis of variance results of regression equation
Note that is significant (P < 0.05) and is very significant (P < 0.01).
As can be seen from the equation, the coefficient of the quadratic term is negative, indicating that the equation has a maximum value. The optimal prediction condition obtained by analyzing the response surface design expert software is as follows: the pH value is 6.5, the temperature is 29.51 ℃, the dosage of the chitosan is 250mg/L, and the predicted protein recovery rate is 85.72 percent.
4 conclusion
According to the model optimization result and the actual conditions, the optimal recovery process of the sarcoplasmic proteins is determined as follows: the pH value is 6.5, the temperature is 30 ℃, the dosage of chitosan is 250mg/L, the flocculation time is 90min, and the recovery rate of the obtained protein under the condition is 85.23%.
Example 2
A method for recovering sarcoplasmic proteins in minced fillet rinsing liquid by acid treatment and chitosan flocculation, which comprises the following steps:
s1: preparing minced fillet rinsing liquid: taking fish meat, adding distilled water into the fish meat for rinsing according to a weight ratio of 1:3 (volume, g/mL), centrifuging, and taking supernate as minced fillet rinsing liquid;
s2: adding a chitosan solution into the minced fillet rinsing liquid, and adjusting the pH value to 7.0;
s3: standing, centrifuging, and precipitating to obtain the co-flocculate of sarcoplasmic protein and a little chitosan.
Preferably, in the step S1, the solid-to-liquid ratio of the fish meat to the distilled water is 1: 10000g,4 ℃,15min.
Preferably, in the step S2, the deacetylation degree of chitosan is 95%, and the addition amount of chitosan in the minced fillet rinsing liquid is 250mg/L.
Preferably, in step S2, the pH is adjusted to 3.0, and then adjusted to 7.0 with NaOH solution.
Preferably, in the step S2, the concentration of the NaOH solution is 3mol/L.
Preferably, in the step S3, after the mixture is placed in a water bath kettle at 20 ℃ and kept still for 90min, the centrifugation conditions are as follows: 4000r/min,10min.
Chemical demand (COD) measurement result
The results of chemical oxygen demand measurement of the rinse solution are shown in Table 3. As can be seen from Table 3, the chemical demand (COD) of the minced fish rinse after flocculation treatment in example 2 was adjusted from the original 674.55 mg.L -1 The temperature is reduced to 179.98 mg.L -1 The COD value removal rate reaches 73.32 percent. Therefore, the acid treatment can improve the protein flocculation effect of the chitosan and can also obviously reduce the chemical oxygen demand in the minced fillet rinsing liquid.
TABLE 3 COD measurement results of surimi rinses
The recovered coprocellate of sarcoplasmic proteins and chitosan is used for making sarcoplasmic proteins electrospun nanofibrous membrane, the application comprising the steps of:
1) Freeze-drying the co-flocculate of the sarcoplasmic protein and the chitosan, dissolving the freeze-dried co-flocculate in hexafluoroisopropanol to prepare solutions with the mass fractions of 2%, 4%, 6%, 8% and 10%, and magnetically stirring the solutions for 24 hours at room temperature to obtain a spinning solution;
2) The spinning solution is subjected to electrostatic spinning at 25 ℃, the humidity is 45%, 20KV electrostatic voltage is used, the spinning speed is 0.8mL/h, and the distance between a spinning nozzle and a receiving plate is 14cm, and an aluminum foil paper is used for receiving a fiber membrane in a rolling mode;
3) And (3) drying the fibrous membrane for 48 hours in vacuum at 40 ℃ to obtain the sarcoplasmic protein electrostatic spinning nanofiber membrane.
Scanning Electron Microscope (SEM) observation of flocculated sarcoplasmic protein electrospun membranes
Example 2 the flocculated sarcoplasmic proteins were used to make sarcoplasmic proteins electrospun nanofiber membranes (flocculated sarcoplasmic proteins electrospun membranes, FL-FSP for short), it can be seen from fig. 7 that at lower concentrations, the solution viscosity was lower and the surface tension was high, the fibers were difficult to form, and the fiber surface was accompanied by a large number of beads, as shown by SEM at 2% and 4%. When the concentration is increased, the viscosity is increased and the surface tension is reduced, so that balanced opposition among the surface tension, the viscosity and the electrostatic repulsion is formed, and thus, flat and uniform fibers are obtained, and no beading is generated on the surface, as shown by a 6% SEM. The solution viscosity was too high, the rheology was poor and spinning was difficult as shown by the SEM at 8% and 10%. Therefore, it was subsequently decided to use a protein dope concentration of 6%.
The results of SEM show that the sarcoplasmic proteins obtained by acid treatment combined with chitosan flocculation can be directly used for electrostatic spinning film formation without separating the chitosan remained in the proteins, thereby reducing the consumption of manpower and material resources.
Contact Angle test results
The flocculation muscle plasma protein electro-spinning membrane (FL-FSP) is prepared by adopting the concentration of 6 percent of protein spinning solution, and is compared with the direct freeze-drying muscle plasma protein electro-spinning membrane (FD-FSP). The preparation method of the muscle plasma protein electro-spinning membrane by direct freeze drying comprises the following steps: the fish meat was rinsed with 1 -1 Centrifuging at 4 deg.C for 15min, freezing the supernatant in-80 deg.C refrigerator, and freeze drying to obtain directly freeze-dried sarcoplasmic protein. Then the protein is dissolved in hexafluoroisopropanol to prepare a spinning solution with the concentration of 8 percent, and the direct freeze-dried sarcoplasmic protein electrospun membrane (FD-FSP) is prepared by electrostatic spinning under the conditions that the temperature is 25 ℃, the humidity is 45 percent, the electrostatic voltage of 20KV is used, the spinning speed is 0.8mL/h, and the distance between a spinning nozzle and a receiving plate is 14 cm.
The contact angle measurement results of the sarcoplasmic protein electrospun membranes in different recovery modes are shown in fig. 8, the contact angle of the directly freeze-dried sarcoplasmic protein electrospun membrane is 97.0 +/-0.9 degrees, and the contact angle of the flocculated sarcoplasmic protein electrospun membrane is 124.1 +/-1.5 degrees, which is probably because after the sarcoplasmic protein is treated by acid, the obtained flocculated sarcoplasmic protein electrospun membrane has rough surface and larger surface energy, so that the contact angle is increased, and the surface hydrophobicity of the fibrous membrane is improved.
Analysis of mechanical Properties
The mechanical properties of the sarcoplasmic protein electrospun fibrous membrane are shown in fig. 9, the elastic modulus and tensile strength of the direct freeze-dried sarcoplasmic protein fibrous membrane are 77.5MPa and 1.0MPa, respectively, while the elastic modulus and tensile strength of the flocculated sarcoplasmic protein fibrous membrane are 29.0MPa and 0.4MPa, respectively, which may be caused by the damage of acid treatment to the protein structure. The elongation at break of the direct freeze-dried and flocculated sarcoplasmic protein fibers was 4.6% and 4.8%, respectively, and the results showed that the acid treatment did not negatively affect the elongation at break of the protein fiber membrane.
Differential Scanning Calorimetry (DSC)
Sarcoplasmic proteins undergo irreversible deformation (contraction) after reaching a certain temperature, the molecules being transformed from an extended helical to a disordered coiled state. The higher the denaturation temperature of the sarcoplasmic proteins, the more stable their internal structure and the better the thermal stability. The thermodynamic behavior of chitosan powder, FD-FSP, FL-FSP fibrous membranes was studied by Differential Scanning Calorimetry (DSC). The DSC of fig. 10 shows that chitosan powder has an endothermic peak at 148.4 ℃, corresponding to the melting point (Tm) of chitosan. The FD-FSP fiber and the FL-FSP fiber show similar endothermic peaks, the melting point of the FD-FSP fiber is 167.0 ℃, the melting point of the FL-FSP fiber membrane is 173.0 ℃, and the endothermic peak value is increased, which shows that the heat stability of the sarcoplasmic protein electrospun membrane is high, and the flocculated sarcoplasmic protein membrane is better than the direct freeze-dried sarcoplasmic protein membrane.
Compared with freeze drying, the flocculation process saves energy and reduces consumption, and the flocculation recovery of the sarcoplasmic proteins reduces the treatment cost and pressure of the minced fillet rinsing wastewater, and solves the problems of recovery and high-value utilization of byproducts in the industrialized production of the minced fillet.
Example 3
A method for recovering sarcoplasmic proteins in minced fillet rinsing liquid by acid treatment and chitosan flocculation, which comprises the following steps:
s1: preparing a minced fillet rinsing liquid: adding distilled water into fish meat for rinsing, centrifuging, and collecting supernatant as minced fillet rinsing liquid;
s2: adding a chitosan solution into the minced fillet rinsing liquid, and adjusting the pH to 6.5;
s3: standing, centrifuging, and precipitating to obtain sarcoplasmic protein and chitosan cochlegmation.
Preferably, in the step S1, the solid-to-liquid ratio of the fish meat to the distilled water is 1: 9000g,4 ℃ for 12min.
Preferably, in the step S2, the deacetylation degree of chitosan is 90%, and the addition amount of chitosan in the minced fillet rinsing liquid is 300mg/L.
Preferably, in step S2, the pH is adjusted to 3.0, and then adjusted to 6.5 with NaOH solution.
Preferably, in the step S2, the concentration of the NaOH solution is 2.5mol/L.
Preferably, in the step S3, the step of placing the mixture into a 20 ℃ water bath kettle to stand is carried out at 15 ℃, and after the standing time is 85min, the centrifugation conditions are as follows: 3500r/min,8min.
The sarcoplasmic proteins prepared by the method are used for manufacturing the sarcoplasmic protein electrostatic spinning nanofiber membrane, and the application comprises the following steps:
1) Freeze-drying the co-flocculate of the sarcoplasmic protein and the chitosan, dissolving the freeze-dried sarcoplasmic protein and the chitosan in hexafluoroisopropanol, and magnetically stirring to obtain a spinning solution;
2) Carrying out electrostatic spinning on the spinning solution, and receiving the fiber membrane by rolling aluminum foil paper;
3) The fiber membrane is dried to remove residual organic solvent.
Preferably, the electrostatic spinning conditions of step 2) are as follows: electrostatic spinning is carried out at 23 ℃, humidity is 40%, 18KV electrostatic voltage is used, spinning speed is 0.6mL/h, and distance between a spinning nozzle and a receiving plate is 12 cm.
Preferably, the drying conditions in step 3) are as follows: naturally drying at room temperature.
Example 4
A method for recovering sarcoplasmic proteins in minced fillet rinsing liquid by acid treatment and chitosan flocculation, which comprises the following steps:
s1: preparing a minced fillet rinsing liquid: adding fish meat into distilled water for rinsing, and centrifuging to obtain supernatant as minced fillet rinsing liquid;
s2: adding a chitosan solution into the minced fillet rinsing liquid, and adjusting the pH to 8.0;
s3: standing, centrifuging, and precipitating to obtain the co-flocculate of sarcoplasmic protein and a little chitosan.
Preferably, in the step S1, the solid-to-liquid ratio of the fish meat to the distilled water is 1: 11000g,4 ℃,18min.
Preferably, in the step S2, the deacetylation degree of chitosan is 90-98%, and the addition amount of chitosan in the minced fillet rinsing liquid is 150mg/L.
Preferably, in step S2, the pH is adjusted to 3.0, and then adjusted to 8.0 with NaOH solution.
Preferably, in the step S2, the concentration of the NaOH solution is 2.5-3.5mol/L.
Preferably, in step S3, the temperature of standing in the 20 ℃ water bath is 25 ℃, and after the standing time is 95min, the centrifugation conditions are as follows: 4500r/min,12min.
The sarcoplasmic proteins prepared by the method are used for manufacturing the sarcoplasmic protein electrostatic spinning nanofiber membrane, and the application comprises the following steps:
1) The precipitation is that sarcoplasmic protein and chitosan are dissolved in hexafluoroisopropanol after the co-flocculate is freeze-dried, and the spinning solution is obtained after magnetic stirring;
2) Carrying out electrostatic spinning on the spinning solution, and receiving the fiber membrane by rolling aluminum foil paper;
3) The fiber membrane was vacuum dried to remove residual organic solvent.
Preferably, the electrostatic spinning conditions of step 2) are as follows: electrostatic spinning is carried out at 27 ℃, the humidity is 0%, 22KV electrostatic voltage is used, the spinning speed is 1.2mL/h, and the distance between a spinning nozzle and a receiving plate is 16 cm.
Preferably, the drying conditions in step 3) are as follows: dried in a vacuum oven at 45 ℃ for 50 hours.
The above-described embodiments are merely illustrative of the preferred embodiments of the present invention, and do not limit the scope of the present invention, and various modifications and improvements of the technical solutions of the present invention can be made by those skilled in the art without departing from the spirit of the present invention, and the technical solutions of the present invention are within the scope of the present invention defined by the claims.
Claims (6)
1. The method for recovering the sarcoplasmic proteins in the minced fillet rinsing liquid by acid treatment and chitosan flocculation is characterized by comprising the following steps:
s1: preparing a minced fillet rinsing liquid: adding distilled water into fish meat for rinsing, centrifuging, and taking supernatant as minced fillet rinsing liquid;
s2: adding a chitosan solution into the minced fillet rinsing liquid, firstly adjusting the pH value to 3, and then adjusting the pH value to 7;
s3: standing, centrifuging, and precipitating to obtain sarcoplasmic protein and chitosan co-flocculate;
in the step S2, the deacetylation degree of the chitosan is 85-98%, and the addition amount of the chitosan in the minced fillet rinsing liquid is 150-400 mg/L;
the coprocellation of the sarcoplasmic protein and the chitosan recovered by the method is used for manufacturing the sarcoplasmic protein electrostatic spinning nanofiber membrane, and comprises the following steps:
1) Freeze-drying the co-flocculate of the sarcoplasmic protein and the chitosan, dissolving the freeze-dried sarcoplasmic protein and the chitosan in hexafluoroisopropanol, and magnetically stirring to obtain a spinning solution;
2) Carrying out electrostatic spinning on the spinning solution, and receiving the fiber membrane by rolling aluminum foil paper;
3) And drying the fiber membrane to remove residual organic solvent to obtain the sarcoplasmic protein electrospun nanofiber membrane.
2. The method of claim 1, wherein in the step S1, the solid-to-liquid ratio of the fish meat to the distilled water is 1 (1.0-3.5) (g/mL), and the centrifugation conditions are as follows: 8000-11000g,4 deg.C, 12-18min.
3. The method of claim 1, wherein in step S2, a NaOH solution is used to adjust the pH to 7, and the concentration of the NaOH solution is 1.0-6.0mol/L.
4. The method according to claim 1, wherein in the step S3, the temperature of the still standing in the water bath is 10-35 ℃, and after the still standing time is 30-300min, the centrifugation conditions are as follows: 3500-4500r/min,8-15min.
5. The method according to claim 1, wherein the step 2) electrospinning conditions are: electrostatic spinning is carried out under the conditions that the temperature is 13-27 ℃, the humidity is 30-65%, the electrostatic voltage is 15-22KV, the spinning speed is 0.4-1.8mL/h, and the distance between a spinning nozzle and a receiving plate is 10-18 cm.
6. The method according to claim 1, wherein the drying conditions of step 3) are: naturally drying at room temperature or drying in a vacuum drying oven at 35-60 deg.C.
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