Fish maw composite enzymolysis-ultrasonic wave synergistic fishy smell and oil removal method

Abstract

According to the fish maw composite enzymolysis-ultrasonic wave synergistic fishy smell and oil removal method, the fish maw is treated through the compound enzyme and ultrasonic waves, fat cell membranes are physically broken through the ultrasonic waves, contact between the compound enzyme and the fish maw is promoted, the compound enzyme can better hydrolyze amino acid at a trimethylamine binding site in the fish maw, and therefore trimethylamine is removed; the removal rate of trimethylamine reaches 98.3%, the grease residue is less than 0.12%, and better oil and fishy smell removal is realized.

Description

Technical Field

[0001] This application belongs to the technical field of aquatic product processing, and particularly relates to a method for removing fishy smell and oil from fish maw by combined enzymatic hydrolysis-ultrasound synergism. Background Art

[0002] Fish maw, also known as fish glue, is the swim bladder of fish and is one of the eight precious ingredients. Because it is rich in high-grade collagen and various trace elements, it is a nourishing and therapeutic food ingredient. However, the fishy smell in fish maw makes many people deterred. Therefore, it is necessary to process fish maw to remove the fishy smell and oil and improve the taste of fish maw.

[0003] Traditional fish maw deodorization processes mostly use acid-base soaking or high-temperature steaming and boiling. Although they can remove the fishy smell of fish maw to a certain extent, they also have the following obvious defects: 1. The risk of chemical residues, such as NaOH causing pH imbalance; 2. High temperature causes collagen denaturation, and the loss rate of collagen > 30%; 3. The treatment cycle is long. Generally, acid-base soaking takes more than 2 hours, and long-term soaking further increases the situation of chemical residues and collagen denaturation loss.

[0004] Therefore, further research is needed on the fish maw deodorization process. Summary of the Invention

[0005] The embodiments of this application provide a method for removing fishy smell and oil from fish maw by combined enzymatic hydrolysis-ultrasound synergism to solve the problems existing in the related technologies. The technical solutions are as follows:

[0006] In a first aspect, the embodiments of this application provide a method for removing fishy smell and oil from fish maw by combined enzymatic hydrolysis-ultrasound synergism, including the following steps:

[0007] Ultrasonically treat the fish maw in a composite enzyme solution, and then raise the temperature for enzymatic hydrolysis to obtain fish maw with removed fishy smell and oil.

[0008] In one embodiment, before ultrasonic treatment, remove the surface impurities of the fish maw with water.

[0009] In one embodiment, the material-liquid ratio of the fish maw to the composite enzyme solution is 1:6-10.

[0010] In one embodiment, the composite enzyme solution is a mixed solution of lipase and flavor protease; wherein the lipase is 500-800 U / g; the flavor protease is 800-1200 U / g.

[0011] In one embodiment, the conditions for ultrasonic treatment are: under 4-10°C, use ultrasonic waves with a frequency of 35-45 kHz and a power density of 40-60 W / L to treat for 20-40 minutes.

[0012] In one embodiment, the temperature is raised to 50 - 60 °C and the enzymatic hydrolysis time is 10 - 20 min.

[0013] In one embodiment, the heating rate is 1 - 2 °C / min.

[0014] In one embodiment, during ultrasonic treatment, mechanical oscillation is applied synchronously. The frequency of the mechanical oscillation is 2 Hz and the amplitude is 5 mm.

[0015] In one embodiment, the pH of the complex enzyme solution is 6.5, which is adjusted by phosphate buffer solution.

[0016] In one embodiment, the complex enzyme solution also contains tea polyphenols, and the concentration of tea polyphenols is 0.1 - 0.3% w / v.

[0017] In one embodiment, after enzymatic hydrolysis, centrifugal separation and water washing are carried out to obtain the fish maw with deodorized and defatted.

[0018] In a second aspect, the embodiments of the present application provide a deoiled and deodorized fish maw, which is prepared by the fish maw complex enzymatic hydrolysis - ultrasonic synergistic deodorization and defatting method described in any one of the above.

[0019] The advantages or beneficial effects in the above technical solutions at least include:

[0020] In the fish maw complex enzymatic hydrolysis - ultrasonic synergistic deodorization and defatting method of the present application, the fish maw is treated with a complex enzyme and ultrasonic waves. The cell membrane of fat cells is physically broken by ultrasonic waves, and the contact between the complex enzyme and the fish maw is promoted. The complex enzyme can better hydrolyze the amino acids at the trimethylamine binding sites in the fish maw, thereby removing trimethylamine and achieving better deodorization and defatting.

[0021] The above summary is only for the purpose of the specification and is not intended to be limiting in any way. In addition to the illustrative aspects, embodiments and features described above, further aspects, embodiments and features of the present application will become apparent from the following detailed description. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 It is a physical diagram of the fish maw for Example 1, Comparative Example 2, and Comparative Example 4. DETAILED DESCRIPTION OF THE EMBODIMENTS

[0023] In the following, only some exemplary embodiments are simply described. As those skilled in the art can recognize, the described embodiments can be modified in various different ways without departing from the spirit or scope of the present application. Therefore, the description is considered to be exemplary in nature and not restrictive.

[0024] In the prior art, when using protease for enzymatic hydrolysis of fish maw, the texture and integrity of the fish maw are often damaged due to the action of protease, and it is enzymatically hydrolyzed into collagen. Therefore, it is not suitable for removing oil and fishy smell from the fish maw itself. However, in this application, a specific compound protease is used in combination with ultrasonic treatment to achieve the removal of oil and fishy smell from the fish maw.

[0025] This application provides a method for synergistically removing fishy smell and oil from fish maw by compound enzymatic hydrolysis - ultrasonic wave, which includes the following steps:

[0026] The fish maw is subjected to ultrasonic treatment in a compound enzyme solution, and then heated for enzymatic hydrolysis to obtain the fish maw with removed fishy smell and oil.

[0027] Ultrasound can produce cavitation. The microjet generated by ultrasonic wave, on the one hand, can promote the contact efficiency between the enzyme and the substrate, which helps to shorten the enzymatic hydrolysis treatment time and reduce the damage of protease to the fish maw; on the other hand, ultrasonic wave can physically break the fat cell membrane and reduce the usage amount of lipase for oil removal.

[0028] As one of the embodiments, the compound enzyme solution is a mixed solution of lipase and flavor protease; wherein the lipase is 500 - 800U / g; the flavor protease is 800 - 1200U / g.

[0029] This application selects a compound enzyme of flavor protease and lipase. The protease therein is flavor protease rather than ordinary neutral / alkaline protease. Flavor protease acts on the fish maw and preferentially hydrolyzes hydrophobic amino acids, such as methionine and phenylalanine at the trimethylamine binding site. Flavor protease cuts off the connection bond between trimethylamine and the carrier protein (such as the Met - TMA complex); it can efficiently remove the fishy smell substance trimethylamine. At the same time, the exopeptidase activity of flavor protease is dominant, and it can start hydrolysis from the end of the fish maw protein peptide chain, avoiding the destruction of the internal structure of collagen.

[0030] As one of the embodiments, before ultrasonic treatment, the impurities on the surface of the fish maw are removed with water. Removing the impurities on the surface of the fish maw, such as obvious fat, etc., can improve the removal of fat by lipase and shorten the enzyme treatment time.

[0031] As one of the embodiments, the material - liquid ratio of the fish maw to the compound enzyme solution is 1:6 - 10. The material - liquid ratio can be 1:6, 1:7, 1:8, 1:9, 1:10; or any value between 1:6 - 10.

[0032] As one of the embodiments, the conditions for ultrasonic treatment are as follows: at 4 - 10 °C, ultrasonic waves with a frequency of 35 - 45 kHz and a power density of 40 - 60 W / L are used for treatment for 20 - 40 min. The micro - jets generated by ultrasonic waves with a frequency of 5 - 45 kHz and a power density of 40 - 60 W / L can increase the contact between the enzyme and the fish maw, and can also physically break the cell membrane of fat, promoting the enzymatic hydrolysis of fat by lipase to remove oil.

[0033] As one of the embodiments, the temperature is raised to 50 - 60 °C, and the enzymatic hydrolysis time is 10 - 20 min. On the one hand, at 50 - 60 °C, especially at 55 °C, the activity of flavor protease is 75% of its maximum activity, ensuring a moderate reaction and achieving the effect of removing fishy smell; while the denaturation temperature of collagen is generally greater than 60 °C, so the enzyme treatment temperature in this application does not exceed 60 °C, and it can also avoid the denaturation of collagen caused by temperature.

[0034] As one of the embodiments, the heating rate is 1 - 2 °C / min. Controlling the heating rate makes the fish maw heat evenly, avoiding too much influence on the texture of the fish maw. And it can slowly activate the enzyme activity to achieve efficient enzymatic hydrolysis.

[0035] As one of the embodiments, during ultrasonic treatment, mechanical oscillation is applied synchronously. The frequency of the mechanical oscillation is 2 Hz, and the amplitude is 5 mm.

[0036] As one of the embodiments, the pH of the composite enzyme solution is 6.5, which is adjusted by phosphate buffer solution.

[0037] As one of the embodiments, the composite enzyme solution also contains tea polyphenols, and the concentration of tea polyphenols is 0.1 - 0.3% w / v. As an antioxidant, tea polyphenols can scavenge free radicals, with an ORAC value ≥ 8000 μmol TE / g, preventing the generation of new fishy smell due to lipid oxidation; and it can also complex with trimethylamine released into the solution by enzymatic hydrolysis to form a complex with a molecular weight > 500 Da, which cannot be perceived by smell, thus further reducing the presence of fishy smell. And the concentration of tea polyphenols is only 0.1 - 0.3% w / v, reducing the color change of the fish maw caused by the use of high - concentration tea polyphenols.

[0038] As one of the embodiments, after enzymatic hydrolysis, centrifugal separation and washing with water are carried out to obtain fish maw with removed fishy smell and oil.

[0039] This application also provides a fish maw with removed oil and fishy smell, which is prepared by the method for synergistic removal of oil and fishy smell by composite enzymatic hydrolysis - ultrasonic wave of the fish maw described in any one of the above.

[0040] The following is further illustrated by specific examples.

[0041] Example 1

[0042] Take 500 g of dried fish maw and rinse the surface impurities with pure water;

[0043] Prepare a complex enzyme solution, lipase (Novozymes 435) 650 U / g of fish maw; flavor protease (Sigma Aldrich 33077) 1000 U / g of fish maw; tea polyphenols (purity ≥ 98%) 0.2% w / v; adjust the pH to 6.5 with phosphate buffer solution;

[0044] Add the fish maw to the complex enzyme solution at a material-liquid ratio of 1:8, with an ultrasonic frequency of 40 kHz and a power density of 50 W / L; ultrasonically treat at 4 °C for 30 min; then gradually increase the temperature to 55 °C and maintain for 15 min; centrifuge and dehydrate at 3000 rpm for 10 min; rinse the fish maw with clear water 3 times, drain, and obtain the defatted and deodorized fish maw.

[0045] Example 2

[0046] Take 500 g of dried fish maw and rinse the surface impurities with pure water;

[0047] Prepare a complex enzyme solution, lipase (Novozymes 435) 500 U / g of fish maw; flavor protease (Sigma Aldrich 33077) 1200 U / g of fish maw; tea polyphenols (purity ≥ 98%) 0.1% w / v; adjust the pH to 6.5 with phosphate buffer solution;

[0048] Add the fish maw to the complex enzyme solution at a material-liquid ratio of 1:10, with an ultrasonic frequency of 45 kHz and a power density of 40 W / L; ultrasonically treat at 10 °C for 20 min; then gradually increase the temperature to 50 °C and maintain for 20 min; centrifuge and dehydrate at 3000 rpm for 10 min; rinse the fish maw with clear water 3 times, drain, and obtain the defatted and deodorized fish maw.

[0049] Example 3

[0050] Take 500 g of dried fish maw and rinse the surface impurities with pure water;

[0051] Prepare a complex enzyme solution, lipase (Novozymes 435) 800 U / g of fish maw; flavor protease (Sigma Aldrich 33077) 800 U / g of fish maw; tea polyphenols (purity ≥ 98%) 0.3% w / v; adjust the pH to 6.5 with phosphate buffer solution;

[0052] Add the fish maw into the complex enzyme solution at a material-liquid ratio of 1:6, with an ultrasonic frequency of 35 kHz and a power density of 60 W / L; ultrasonically treat it at a temperature of 7 °C for 40 min; then gradually increase the temperature to 60 °C and maintain for 10 min; centrifuge and dehydrate at 3000 rpm for 10 min; rinse the fish maw with clear water 3 times, drain, and obtain the defatted and deodorized fish maw.

[0053] Control Example 1

[0054] Take 500 g of dried fish maw and rinse the surface impurities with pure water;

[0055] Add the fish maw into the 1% citric acid solution at a material-liquid ratio of 1:8, with an ultrasonic frequency of 40 kHz and a power density of 50 W / L; ultrasonically treat it at a temperature of 4 °C for 30 min; then gradually increase the temperature to 55 °C and maintain for 90 min; centrifuge and dehydrate at 3000 rpm for 10 min; rinse the fish maw with clear water 3 times, drain, and obtain the defatted and deodorized fish maw.

[0056] Control Example 2

[0057] Take 500 g of dried fish maw and rinse the surface impurities with pure water;

[0058] Prepare the complex enzyme solution, with lipase 650 U / g of fish maw; neutral protease 1000 U / g of fish maw; tea polyphenol (purity ≥ 98%) 0.2% w / v; adjust the pH to 6.5 with phosphate buffer solution;

[0059] Add the fish maw into the complex enzyme solution at a material-liquid ratio of 1:8, with an ultrasonic frequency of 40 kHz and a power density of 50 W / L; ultrasonically treat it at a temperature of 4 °C for 30 min; then gradually increase the temperature to 55 °C and maintain for 15 min; centrifuge and dehydrate at 3000 rpm for 10 min; rinse the fish maw with clear water 3 times, drain, and obtain the defatted and deodorized fish maw.

[0060] Control Example 3

[0061] Take 500 g of dried fish maw and rinse the surface impurities with pure water;

[0062] Prepare the complex enzyme solution, with lipase 650 U / g of fish maw; flavor protease 1000 U / g of fish maw; tea polyphenol (purity ≥ 98%) 0.2% w / v; adjust the pH to 6.5 with phosphate buffer solution;

[0063] Add the fish maw into the complex enzyme solution at a material-liquid ratio of 1:8, with an ultrasonic frequency of 40 kHz and a power density of 30 W / L; ultrasonically treat it at a temperature of 4 °C for 30 min; then gradually increase the temperature to 55 °C and maintain for 15 min; centrifuge and dehydrate at 3000 rpm for 10 min; rinse the fish maw with clear water 3 times, drain, and obtain the defatted and deodorized fish maw.

[0064] Control Example 4

[0065] Take 500 g of dried fish maw and rinse the surface impurities with pure water;

[0066] Prepare a compound enzyme solution, with lipase 650 U / g of fish maw; flavor protease 1000 U / g of fish maw; tea polyphenols (purity ≥ 98%) 0.5% w / v; adjust the pH to 6.5 with phosphate buffer solution;

[0067] Add the fish maw to the compound enzyme solution according to a solid-liquid ratio of 1:8, with an ultrasonic frequency of 40 kHz and a power density of 50 W / L; perform ultrasonic treatment at a temperature of 4 °C for 30 min; then gradually increase the temperature to 55 °C and maintain for 15 min; centrifuge for dehydration at 3000 rpm for 10 min; rinse the fish maw with clear water 3 times, drain, and obtain the defatted and deodorized fish maw.

[0068] Determine the relative content of trimethylamine in the fish maw by GC-MS. Sample preparation: Accurately weigh the sample into a centrifuge tube, add trichloroacetic acid solution for treatment, and centrifuge after treatment; filter the supernatant through absorbent cotton, and collect the filtrate in a volumetric flask. The residue is extracted once with trichloroacetic acid solution respectively, combine the 3 filtrates, and dilute and make up the volume with 5% trichloroacetic acid solution. Accurately transfer the above sample solution into a headspace vial, seal the cap, and accurately inject sodium hydroxide solution from the edge of the septum with a syringe, and determine according to the instrument working conditions; calculate the content of trimethylamine in the fish maw before and after treatment, and the results are shown in Table 1.

[0069] Detect the hardness of the fish maw in Example 1 and Comparative Examples 1-4 by a texture analyzer, and the results are shown in Table 1.

[0070] Detect the oil content in the fish maw of Example 1 and Comparative Examples 1-4, and the results are shown in Table 1.

[0071] Detection method for oil residue

[0072] 1. Detection standard

[0073] According to the acid hydrolysis method in "GB 5009.6-2016 National Food Safety Standard Determination of Fat in Foods", perform quantitative analysis on the bound fat in the fish maw.

[0074] 2. Experimental steps

[0075] 2.1 Sample pretreatment: Take 10.0 g of the treated fish maw sample, freeze-dry and crush it to 80 mesh; add 10 mL of 8 mol / L HCl solution, hydrolyze in a boiling water bath for 2 hours; cool and filter, wash the filter residue with distilled water until neutral, and dry at 60 °C to constant weight.

[0076] 2.2 Fat extraction: Place the dried sample in a Soxhlet extractor, add 50 mL of petroleum ether (boiling range 30-60 °C); the water bath temperature is 60 °C, and reflux extraction is carried out for 6 hours;

[0077] 2.3 Weighing and calculation: Recover petroleum ether, and dry the fat bottle in an oven at 105 °C until constant weight.

[0078] Residual oil content (%) = (mass of extracted fat / mass of sample) × 100%.

[0079] Detect the collagen content in the fish maw of Example 1 and Comparative Examples 1-4, and calculate the collagen retention rate. The results are shown in Table 1.

[0080] Method for detecting collagen retention rate

[0081] 1. Detection standard

[0082] Adopt the spectrophotometry method in "GB 5009.124-2016 National Food Safety Standard Determination of Hydroxyproline in Foods", and combine SDS-PAGE electrophoresis to verify the integrity of collagen.

[0083] 2. Determination of hydroxyproline content

[0084] 2.1 Sample hydrolysis: Take 1.0 g of the fish maw sample before and after treatment, add 10 mL of 6 mol / L HCl solution; hydrolyze at 110 °C for 24 hours, cool and make up the volume to 100 mL, and filter for standby.

[0085] 2.2 Color reaction: Take 1 mL of the filtrate, add 1 mL of chloramine T solution (1.4 g / L), oxidize at room temperature for 10 minutes; add 2 mL of p-dimethylaminobenzaldehyde solution (10% perchloric acid solution), and develop color in a water bath at 60 °C for 20 minutes.

[0086] 2.3 Absorbance measurement: Measure the absorbance at a wavelength of 558 nm, and calculate the hydroxyproline content according to the standard curve.

[0087] 2.4 Retention rate calculation: Collagen retention rate (%) = (hydroxyproline content after treatment / hydroxyproline content before treatment) × 100%.

[0088] Compare the appearances of the fish maw in Example 1 and Comparative Examples 2 and 4. The appearance diagrams are as Figure 1 shown, where from left to right are Example 1, Comparative Example 2, and Comparative Example 4. Table 1

[0089]

[0090] From Figure 1It can be seen that the appearance of the fish maw in Example 1 is intact and not much different from that of the fish maw before treatment. However, the fish maw in Comparative Example 2 is basically enzymatically hydrolyzed, and the overall structure of the fish maw is completely destroyed. Moreover, the brown color of the fish maw in Comparative Example 4 is significantly deepened. After detection, the L* value decreases by 8.2, while the color change of the fish maw in Example 1 of the present application is indistinguishable to the naked eye.

[0091] From the data in Table 1, it can be known that under the conditions of the examples of the present application, the content of the fishy smell substance trimethylamine is reduced by 98.3%. For the traditional acid treatment, after treatment for up to 120 minutes, only 72.5% of trimethylamine is removed. Under the action of neutral protease in Comparative Example 2, the fish maw is severely enzymatically hydrolyzed. In Comparative Example 3, the ultrasonic power density is changed, and the removal rate of trimethylamine decreases significantly, indicating that the ultrasonic power has a great influence on the removal of trimethylamine and also indicating the necessity of ultrasonic treatment. In Comparative Example 4, more tea polyphenols are used, which not only increases the treatment cost, but also the excessive tea polyphenols cause the color browning of the fish maw to be aggravated, affecting its appearance.

[0092] In summary, the present application uses the combined treatment of complex enzyme and ultrasonic treatment for the fish maw. By ultrasonic physical fragmentation of the cell membrane of fat cells and promoting the contact between the complex enzyme and the fish maw, the complex enzyme can better hydrolyze the amino acids at the trimethylamine binding sites in the fish maw, thereby removing trimethylamine and achieving better degreasing and deodorization.

[0093] In the description of this specification, the descriptions with reference to terms such as "one embodiment", "some embodiments", "examples", "specific examples", or "some examples" etc. mean that the specific features, structures, materials, or characteristics described in connection with the embodiment or example are included in at least one embodiment or example of the present application. Moreover, the specific features, structures, materials, or characteristics described can be combined in a suitable manner in any one or more embodiments or examples. In addition, without contradiction, those skilled in the art can combine and combine the different embodiments or examples described in this specification and the features of different embodiments or examples.

[0094] In addition, the terms "first" and "second" are only used for descriptive purposes and cannot be understood as indicating or implying relative importance or implicitly specifying the quantity of the indicated technical features. Thus, the features defined with "first" and "second" can explicitly or implicitly include at least one of the features. In the description of the present application, "a plurality" means two or more unless otherwise specifically defined.

[0095] The above is only the specific implementation manner of the present application, but the protection scope of the present application is not limited thereto. Any person skilled in the art within the technical scope disclosed by the present application can easily think of various changes or substitutions, and these should all be covered within the protection scope of the present application. Therefore, the protection scope of the present application should be subject to the protection scope of the claims.

Claims

1. A method for removing fishy smell and oil by the synergistic effect of compound enzymatic hydrolysis and ultrasonic treatment of fish maw, characterized in that, It includes the following steps: The fish maw is ultrasonically treated in a complex enzyme solution and then heated for enzymatic hydrolysis to obtain a fish maw with removed fishy smell and oil.

2. The method for synergistically removing fishy smell and oil from fish maw by complex enzymatic hydrolysis - ultrasonic wave according to claim 1, characterized in that Before ultrasonic treatment, the impurities on the surface of the fish maw are removed with water.

3. The method for synergistically removing fishy smell and oil from fish maw by complex enzymatic hydrolysis - ultrasonic wave according to claim 1, characterized in that The material - liquid ratio of the fish maw to the complex enzyme solution is 1:6 - 10; The complex enzyme solution is a mixed solution of lipase and flavor protease; wherein the lipase is 500 - 800U / g; the flavor protease is 800 - 1200U / g.

4. The method for synergistically removing fishy smell and oil from fish maw by complex enzymatic hydrolysis - ultrasonic wave according to claim 1, characterized in that The conditions of ultrasonic treatment are: under the temperature of 4 - 10°C, using ultrasonic waves with a frequency of 35 - 45kHz and a power density of 40 - 60W / L for treatment for 20 - 40min.

5. The method for synergistically removing fishy smell and oil from fish maw by complex enzymatic hydrolysis - ultrasonic wave according to claim 1, characterized in that Heat up to the temperature of 50 - 60°C, the enzymatic hydrolysis time is 10 - 20min; the heating rate is 1 - 2°C / min.

6. The method for synergistically removing fishy smell and oil from fish maw by complex enzymatic hydrolysis - ultrasonic wave according to claim 1, characterized in that During ultrasonic treatment, mechanical oscillation is synchronously applied, and the frequency of the mechanical oscillation is 2Hz and the amplitude is 5mm.

7. The method for synergistically removing fishy smell and oil from fish maw by complex enzymatic hydrolysis - ultrasonic wave according to claim 1, characterized in that The pH of the complex enzyme solution is 6.5, which is adjusted by phosphate buffer solution.

8. The method for synergistically removing fishy smell and oil from fish maw by complex enzymatic hydrolysis - ultrasonic wave according to claim 1, characterized in that The complex enzyme solution also contains tea polyphenols, and the concentration of tea polyphenols is 0.1 - 0.3% w / v.

9. The method for synergistically removing fishy smell and oil from fish maw by complex enzymatic hydrolysis - ultrasonic wave according to claim 1, characterized in that After the enzymatic hydrolysis is completed, centrifugal separation and water washing are carried out to obtain a fish maw with removed fishy smell and oil.

10. A fish maw for removing oil and fishy smell, characterized in that, Prepared by the method for synergistically removing fishy smell and oil from fish maw by complex enzymatic hydrolysis - ultrasonic wave according to any one of claims 1 - 9.

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