CN111838586A - Enzymolysis method of mussel cooking liquor - Google Patents
Enzymolysis method of mussel cooking liquor Download PDFInfo
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- 238000010438 heat treatment Methods 0.000 claims description 5
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L17/00—Food-from-the-sea products; Fish products; Fish meal; Fish-egg substitutes; Preparation or treatment thereof
- A23L17/50—Molluscs
-
- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L17/00—Food-from-the-sea products; Fish products; Fish meal; Fish-egg substitutes; Preparation or treatment thereof
- A23L17/65—Addition of, or treatment with, microorganisms or enzymes
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23L—FOODS, FOODSTUFFS, OR NON-ALCOHOLIC BEVERAGES, NOT COVERED BY SUBCLASSES A21D OR A23B-A23J; THEIR PREPARATION OR TREATMENT, e.g. COOKING, MODIFICATION OF NUTRITIVE QUALITIES, PHYSICAL TREATMENT; PRESERVATION OF FOODS OR FOODSTUFFS, IN GENERAL
- A23L27/00—Spices; Flavouring agents or condiments; Artificial sweetening agents; Table salts; Dietetic salt substitutes; Preparation or treatment thereof
- A23L27/10—Natural spices, flavouring agents or condiments; Extracts thereof
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- G—PHYSICS
- G06—COMPUTING; CALCULATING OR COUNTING
- G06F—ELECTRIC DIGITAL DATA PROCESSING
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Abstract
The invention discloses a method for carrying out enzymolysis on mussel cooking liquor, which comprises the following steps: concentrating the mussel decoction to obtain a mussel concentrate; adding 0.3-0.7 g of flavourzyme into 100ml of mussel concentrated solution, adjusting the pH value to 6.5-7.5, and carrying out enzymolysis for 2-4 h at the enzymolysis temperature of 45-55 ℃. By adopting the method, the enzymatic hydrolysate of the mussel cooking liquor which has unique flavor and can be used as a seasoning can be obtained.
Description
Technical Field
The invention relates to an enzymolysis method of an aquatic product cooking liquor, in particular to an enzymolysis process for optimizing mussel cooking liquor by using a response surface method, and belongs to the field of seasonings.
Background
The mussel has high yield and strong growing power, is the main cultured mussel in China, has the yield more than half of the world yield, and is frozen or dried as the main processing means at present. In the mussel processing process, about 1.5 tons of mussel cooking liquor is produced for every 1 ton of mussel product. The mussel cooking liquid contains a large amount of nutrient components, and if the mussel cooking liquid is directly discharged into a water body, the environment is polluted and resources are wasted. Therefore, the natural mussel seasoning is developed by utilizing the cooking liquor in the mussel processing process, the problem of resource waste and environmental pollution is solved, the added value of the mussel processing product is increased, and good social and economic benefits are generated.
2017100382495 entitled "Process for enzymatic purification of mussel meat and cooking liquor", comprises: desalting water for cooking mussels to obtain a concentrated solution, washing mussels, collecting meat, crushing to obtain a uniform slurry, mixing the concentrated solution and the uniform slurry of mussel meat to obtain a mixture, and adopting compound protease; heating the mixture for enzymolysis to obtain an enzymolysis liquid, filtering by a ceramic membrane to obtain a clear liquid, separating the clear liquid by a membrane to obtain a trapped liquid and a permeate liquid, wherein the trapped liquid is the mussel polysaccharide liquid; deodorizing the permeate, decolorizing, and passing through a column to obtain a fine filtrate; concentrating the fine filtrate under reduced pressure by evaporation to obtain concentrated solution, spray drying the concentrated solution to obtain mussel oligopeptide powder, adding ethanol into the trapped solution, stirring, standing, collecting precipitate, and drying to obtain crude mussel polysaccharide.
Disclosure of Invention
The invention aims to solve the problem of providing a method for carrying out enzymolysis on mussel cooking liquor, and an enzymolysis product obtained by adopting the method has unique flavor and can be used as a seasoning.
In order to solve the technical problems, the invention provides a method for enzymolysis of mussel cooking liquor, which comprises the following steps: concentrating the mussel decoction to obtain a mussel concentrate; adding 0.3-0.7 g (preferably 0.5-0.7 g) of flavourzyme into 100ml of mussel concentrated solution, adjusting the pH value to 6.5-7.5, and carrying out enzymolysis for 2-4 h at the enzymolysis temperature of 45-55 ℃.
The improvement of the method for the enzymolysis of the mussel cooking liquor of the invention comprises the following steps: adding 0.63g of flavourzyme into 100ml of mussel concentrated solution, adjusting pH to 6.9, and carrying out enzymolysis at 51 ℃ for 3.3 h.
The method for the enzymolysis of the mussel cooking liquor is further improved as follows: and heating and concentrating the mussel cooking liquor at 60 ℃ to 9-11% of the original volume to obtain the mussel concentrated liquor.
The method for the enzymolysis of the mussel cooking liquor is further improved as follows: and (3) after the enzymolysis time is up, inactivating enzyme (inactivating enzyme in boiling water bath for 6 min), cooling to room temperature, centrifuging, and centrifuging to obtain supernate which is enzymolysis liquid.
In the invention process, a response surface method is utilized to optimize the enzymatic hydrolysis process of the mussel cooking liquor.
The method comprises the following steps of firstly, optimizing an enzymolysis process of mussel cooking liquor by a response surface method:
1) and enzymolysis:
heating and concentrating the mussel decoction at 60 deg.C for 24 hr to obtain concentrated mussel solution (10% of the original volume).
Putting 4 conical bottles into 100ml of mussel concentrated solution, adding flavourzyme, trypsin, papain and compound protease into the mussel concentrated solution respectively, and then carrying out enzymolysis on mussel cooking liquor under the optimal enzymolysis conditions of the four proteases; as described in table 1.
2) And centrifuging: inactivating enzyme in boiling water bath for 6min after enzymolysis is finished, cooling to room temperature after enzyme inactivation, and centrifuging for 10min at 4000 r/min;
3) And enzyme species screening: testing the centrifugate obtained in the step 2), determining the hydrolysis degree of the centrifugate, and selecting the flavourzyme as a subsequent enzyme, wherein the measured hydrolysis degree of the flavourzyme is the highest;
4) the method comprises the following steps of (1) determining the value ranges of four influencing factors by carrying out single-factor test on flavourzyme, optimizing the temperature, pH, time and enzyme addition amount on a four-factor three-level by adopting a response surface method, and optimizing the enzymatic hydrolysis process of mussel cooking liquor by taking the hydrolysis degree of the mussel cooking liquor as a response value and applying Design Expert 10.0 software;
TABLE 1 enzymatic hydrolysis conditions for four enzyme preparations
Note: the enzyme addition amount of 0.3% means that 0.3g of enzyme is added to 100ml of mussel concentrate, and the rest is the same.
TABLE 2 response surface design test factor levels
5) And selecting samples with different degrees of hydrolysis from the response surface test and selecting the sample with the optimal experimental condition for flavor analysis.
Second, method for measuring degree of hydrolysis
And (3) determining the degree of hydrolysis: refer to GBT5009.39-2003 formaldehyde potentiometric titration.
Degree of Hydrolysis (DH) ═ content of amino acid nitrogen in the centrate/total nitrogen content in the centrate.
Third, analysis of experimental results
(1) Effect of enzyme addition on the degree of hydrolysis
The enzyme is added in an amount of 0.1%, 0.3%, 0.5%, 0.7%, 0.9%, and the hydrolysis temperature is fixed at 50 deg.C for 3 hr, and pH is 7.0.
As can be seen from FIG. 1, the degree of hydrolysis increases with the increase of the protease content, and tends to be stable when the amount of the added enzyme is 0.5%, because the concentration of the enzyme is gradually saturated by the substrate, and the added enzyme is self-hydrolyzed by adding too much protease, which may have a certain effect on the substrate, and the amount of the added enzyme is preferably selected from 0.3% to 0.7%.
(2) Influence of enzymolysis time on hydrolysis degree
The enzymolysis time is controlled to be 1h, 2h, 3h, 4h and 5h, the fixed hydrolysis temperature is 50 ℃, the pH value is 7.0, and the addition amount of enzyme is 0.5%.
As can be seen from FIG. 2, the hydrolysis degree shows a trend of increasing first and then decreasing as the enzymolysis time increases, and reaches a maximum value at 3h of hydrolysis. Therefore, the enzymolysis time is selected to be more suitable for 2-4 h.
(3) Influence of pH on the degree of hydrolysis
The pH is controlled at 6, 6.5, 7, 7.5 and 8, the hydrolysis temperature is fixed at 50 ℃, the addition of enzyme is 0.5 percent, and the time is 3 hours.
From FIG. 3, it can be seen that the degree of hydrolysis is highest at pH7, and that the degree of hydrolysis above or below 7 is lower than that at a higher value because each enzyme has its optimum pH, and the dissociation groups of the enzyme and the substrate protein are in a state of dissociation where they are easily bound and converted into the product only under appropriate conditions, otherwise the activity of the enzyme is inhibited or even inactivated. Therefore, in the enzymolysis process, the pH is selected to be 6.5-7.5.
(4) Influence of temperature on the degree of hydrolysis
The temperature is controlled at 40 deg.C, 45 deg.C, 50 deg.C, 55 deg.C, 60 deg.C, the addition amount of immobilized enzyme is 0.5%, the time is 3h, and the pH is 7.0.
As can be seen from FIG. 4, when the temperature is 40-50 ℃, the hydrolysis degree is always in an increasing state, because the temperature is in a leading position before the temperature is lower than the optimum temperature, the molecular motion is severe along with the increase of the temperature, the contact chance of the enzyme and the substrate is increased, the reaction rate is accelerated, and the hydrolysis degree is increased; the temperature reaches the highest value at 50 ℃, and tends to be stable at 50-55 ℃, but the temperature is obviously reduced at the temperature higher than 55 ℃, because the enzyme is an active protein, when the temperature is too high, the structure of the enzyme is changed, the activity of the enzyme is reduced, and the reaction rate is reduced. Therefore, the enzymolysis temperature is preferably selected to be 45-55 ℃ in the analysis.
Fourth, response surface optimization design
The method is characterized in that Design-expert8.0.6 software is utilized, four-factor three-level experimental Design is carried out according to Box-Benhnken Design principles, four factors, namely enzymolysis temperature (A), enzymolysis pH (B), enzymolysis time (C) and enzyme addition amount (D), are used as independent variables, the hydrolysis degree (Y) of the mussel cooking liquor is measured to serve as a response value, response surface analysis experiments are carried out, and experimental analysis schemes and results are shown in table 3.
TABLE 3 Experimental analysis protocol and results for response surface
Regression equation analysis of variance
Taking the enzyme adding amount, the enzymolysis time, the enzymolysis temperature and the enzymolysis pH as independent variables, taking the hydrolysis degree of the cooking liquor as a response value, and carrying out quadratic polynomial model fitting of nonlinear regression by using Design Expert 8.0.6 software, wherein the predicted model is as follows: y ═ 13.86+0.13X1-0.17X2+0.088X3+0.20X4+0.11X1X2-0.027X1X3+0.045X1X4+7.500E-003X2X3-0.11X2X4-0.030X3X4-0.22X12-0.30X22-0.22X32-0.18X42。
Analysis of variance and comparison of significance are shown in table 4.
TABLE 4 analysis of variance of regression model of degree of hydrolysis
As can be seen from FIG. 4, model P<0.0001, which shows that the response surface regression model is extremely remarkable; mismatching term P-0.4624>0.05 is not significant, which shows that the regression equation can better fit a real response surface. From the influence of four factors of X1, X2, X3 and X4 on the hydrolysis degree, the factors of X1, X2, X3, X4 and X12、X22、X32、X42Has obvious effect on the result (P)<0.05), X1X2, X2X3, X2X4, X3X4 did not significantly affect the results (P)>0.05). The primary and secondary influence of the factors can be judged by comparing the absolute value of the first-order coefficient of the equation, and the influence factors of the experiment are sorted into X3>X4>X2>X1。
And fixing every two of the four factors in the model, wherein two of the four factors are fixed at the level of 0 to obtain a submodel of the interaction of the other two factors on the hydrolysis degree Y, and the graphs in the figures 5-10 are response surface graphs of the influence of every two of the four factors on the hydrolysis degree.
Best condition verification test
The optimal combination of the four optimized influencing factors obtained by response surface analysis is that the enzymolysis temperature is 51.08 ℃, the pH value is 6.87, the enzymolysis time is 3.26h, the enzyme adding amount is 0.63%, and the predicted hydrolysis degree is 13.9805%. In order to test the accuracy of model prediction, experiments are carried out under the conditions of the optimal enzymolysis temperature of 51 ℃, pH6.9, enzymolysis time of 3.3h and enzyme addition amount of 0.63 percent, the measured hydrolysis degree is 14.01 percent and is basically close to the predicted value, the predicted value and the true value have certain fitness, and the feasibility of the model is further verified.
And fifthly, analyzing the flavor substances of the enzymolysis liquid by adopting a gas chromatography-mass spectrometry combined technology.
Namely, by using Design Expert 8.0.6 software and adopting a Box-Behnken response surface Design method, on the basis of a single-factor experiment, four factors of enzymolysis time, enzymolysis temperature, enzymolysis pH and enzyme dosage are selected as corresponding variables, a single-factor experiment optimal point is taken as a center, a horizontal value is taken from the upper part and the lower part of the optimal point as the level of a response surface, the hydrolysis degree is taken as a response value, polynomial fitting regression is carried out on the corresponding variables to obtain a regression equation, and the regression equation is calculated to obtain the optimal enzymolysis condition.
By adopting the method, the mussel cooking liquor enzymatic hydrolysate with unique flavor can be obtained and can be used as a seasoning.
Drawings
The following describes embodiments of the present invention in further detail with reference to the accompanying drawings.
FIG. 1 is a graph showing the effect of the amount of enzyme added on the degree of hydrolysis;
FIG. 2 is a graph showing the effect of enzymatic hydrolysis time on the degree of hydrolysis;
FIG. 3 is a graph showing the effect of enzymatic pH on the degree of hydrolysis;
FIG. 4 is a graph showing the effect of enzymatic hydrolysis temperature on the degree of hydrolysis;
FIG. 5 is a graph of the response of the interaction of enzymatic temperature and enzymatic pH;
FIG. 6 is a graph of the response of the interaction of enzymatic hydrolysis temperature and enzymatic hydrolysis time;
FIG. 7 is a graph of the response of the interaction of enzyme hydrolysis temperature and enzyme dosage;
FIG. 8 is a graph of the response of the interaction of enzymatic pH and enzymatic time;
FIG. 9 is a graph of the response of the interaction of enzymatic pH and enzyme dosage;
FIG. 10 is a graph showing the response of the interaction between the time of enzymatic hydrolysis and the amount of enzyme added.
Detailed Description
The invention will be further described with reference to specific examples, but the scope of the invention is not limited thereto:
the enzyme activity of the flavourzyme is about 8 multiplied by 104U/g, flavourzyme produced by Guangzhou Huaqi biology, Inc. can be selected.
Example 1-9, a method for mussel cooking liquor enzymolysis, sequentially performing the following steps:
1) And heating the fresh mussel cooking liquor at 60 ℃ for 24 hours, thereby concentrating to 10% of the original volume to obtain the mussel concentrated liquor.
2) Adding flavourzyme into 100ml of mussel concentrated solution, adjusting the pH value, and carrying out enzymolysis at a specific enzymolysis temperature for a corresponding time. Enzyme dosage, enzymolysis temperature, enzymolysis time and pH are specifically shown in the following table 5.
0.3% of the total amount of the enzyme, namely, 0.3g of flavourzyme is added into 100ml of mussel concentrated solution, and the rest is the same.
3) After the enzymolysis time is up, inactivating the enzyme in a boiling water bath for 6min to inactivate the enzyme, then cooling to room temperature, and centrifuging for 10min at 4000 r/min; the supernatant obtained by centrifugation is the enzymolysis liquid.
The flavor components and contents of the enzymatic hydrolysate were analyzed, and the results are shown in table 6 below. The degree of hydrolysis is shown in Table 5.
TABLE 5
| Amount of enzyme added (%) | Temperature (. degree.C.) | Time (h) | pH | Degree of |
|
| 1 | 0.3 | 45 | 3 | 7 | 13.11 |
| 2 | 0.5 | 50 | 3 | 7 | 13.98 |
| 3 | 0.7 | 50 | 3 | 6.5 | 13.79 |
| 4 | 0.5 | 55 | 2 | 7 | 13.50 |
| 5 | 0.5 | 50 | 4 | 7 | 13.98 |
| 6 | 0.3 | 45 | 3 | 6.5 | 13.11 |
| 7 | 0.5 | 50 | 2 | 7.5 | 13.10 |
| 8 | 0.5 | 55 | 3 | 6.5 | 13.42 |
| 9 | 0.63 | 51 | 3.3 | 6.9 | 14.01 |
TABLE 6
The number of useful peaks detected by the sample No. 9 is the highest in the aspect of flavor, and the contents of aldehyde compounds and pyrazine compounds which play an important role in the flavor of the mussel cooking liquid are higher than those of other samples; from the viewpoint of the degree of hydrolysis, the degree of hydrolysis of sample No. 9 measured by formaldehyde titration was also the highest, and therefore sample No. 9 was considered to be the best sample.
Comparative example 1, the flavourzyme in example 9 is changed into the compound protease as 2017100382495, and the hydrolysis is carried out under the optimal enzymolysis condition; the rest is equivalent to example 9.
The final results were: the total content of pyrazine compounds is 51.1%, and the total content of aldehyde compounds is 3.2%.
Comparative example 2-1, the preparation of the mussel concentrate of example 9 was modified to:
the fresh mussel cooking liquor is heated at 60 ℃ to be concentrated to 20% of the original volume, so as to obtain mussel concentrated liquor A.
Replacing 100ml of mussel concentrate with 200ml of mussel concentrate a, thereby achieving the same effective amount; the amount of flavourzyme used was the same as in example 9; the rest is equivalent to example 9.
The final results were: the degree of hydrolysis is 13.56%, the total content of pyrazine compounds is 54%, and the total content of aldehyde compounds is only 4.7%.
Finally, it is also noted that the above-mentioned lists merely illustrate a few specific embodiments of the invention. It is obvious that the invention is not limited to the above embodiments, but that many variations are possible. All modifications which can be derived or suggested by a person skilled in the art from the disclosure of the present invention are to be considered within the scope of the invention.
Claims (4)
1. The method for enzymolysis of mussel cooking liquor is characterized by comprising the following steps:
concentrating the mussel decoction to obtain a mussel concentrate;
adding 0.3-0.7 g of flavourzyme into 100ml of mussel concentrated solution, adjusting the pH value to 6.5-7.5, and carrying out enzymolysis for 2-4 h at the enzymolysis temperature of 45-55 ℃.
2. The method for enzymatic hydrolysis of mussel cooking liquor according to claim 1, wherein:
adding 0.63g of flavourzyme into 100ml of mussel concentrated solution, adjusting pH to 6.9, and carrying out enzymolysis at 51 ℃ for 3.3 h.
3. The method for enzymatic hydrolysis of mussel cooking liquor according to claim 1 or 2, wherein:
and heating and concentrating the mussel cooking liquor at 60 ℃ to 9-11% of the original volume to obtain the mussel concentrated liquor.
4. The method for enzymatic hydrolysis of mussel cooking liquor according to claim 3, wherein:
and (4) after the enzymolysis time is up, inactivating enzyme, cooling to room temperature, centrifuging, and obtaining supernate as enzymolysis liquid.
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| CN202010612222.4A CN111838586A (en) | 2020-06-30 | 2020-06-30 | Enzymolysis method of mussel cooking liquor |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114081135A (en) * | 2021-11-23 | 2022-02-25 | 浙江工商大学 | Mussel soybean paste and preparation method thereof |
| CN116035190A (en) * | 2022-12-30 | 2023-05-02 | 浙江工业大学 | Mussel flavor base material produced by recycling protein from mussel processing byproducts and application |
-
2020
- 2020-06-30 CN CN202010612222.4A patent/CN111838586A/en active Pending
Non-Patent Citations (1)
| Title |
|---|
| 陶美洁: "贻贝蒸煮液浓缩及风味改良研究", 《中国优秀博硕士学位论文全文数据库(硕士)工程科技Ⅰ辑》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN114081135A (en) * | 2021-11-23 | 2022-02-25 | 浙江工商大学 | Mussel soybean paste and preparation method thereof |
| CN116035190A (en) * | 2022-12-30 | 2023-05-02 | 浙江工业大学 | Mussel flavor base material produced by recycling protein from mussel processing byproducts and application |
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