CN116035182B - Hypoallergenic cod surimi fermented by lactobacillus helveticus and preparation method thereof - Google Patents
Hypoallergenic cod surimi fermented by lactobacillus helveticus and preparation method thereof Download PDFInfo
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Classifications
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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
- A23L17/65—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
- A23L17/00—Food-from-the-sea products; Fish products; Fish meal; Fish-egg substitutes; Preparation or treatment thereof
- A23L17/70—Comminuted, e.g. emulsified, fish products; Processed products therefrom such as pastes, reformed or compressed products
-
- 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
- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
- A23L33/135—Bacteria or derivatives thereof, e.g. probiotics
-
- 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
- A23L5/00—Preparation or treatment of foods or foodstuffs, in general; Food or foodstuffs obtained thereby; Materials therefor
- A23L5/20—Removal of unwanted matter, e.g. deodorisation or detoxification
- A23L5/28—Removal of unwanted matter, e.g. deodorisation or detoxification using microorganisms
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23V—INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION
- A23V2002/00—Food compositions, function of food ingredients or processes for food or foodstuffs
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- Life Sciences & Earth Sciences (AREA)
- Polymers & Plastics (AREA)
- Health & Medical Sciences (AREA)
- Nutrition Science (AREA)
- Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Food Science & Technology (AREA)
- Marine Sciences & Fisheries (AREA)
- Zoology (AREA)
- Microbiology (AREA)
- Mycology (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
Abstract
The invention relates to the technical field of food processing, in particular to hypoallergenic cod surimi fermented by lactobacillus helveticus and a preparation method thereof. A preparation method of hypoallergenic cod minced fillet fermented by Lactobacillus helveticus comprises mixing and fermenting the cod minced fillet, salt, glucose and Lactobacillus helveticus; the lactobacillus helveticus is one or more of lactobacillus helveticus Lh187926, lactobacillus helveticus Lh191404 and lactobacillus helveticus Lh189793. According to the invention, the sensitization of the cod chyme allergic protein is reduced by adopting the processing modes of fermentation of different lactobacillus helveticus strains, the optimal strain is preferably obtained, the allergen reduction efficiency is improved, and a better reduction effect is obtained; the prepared hypoallergenic cod surimi is delicious in taste and rich in nutrition, can provide various essential amino acids, colors, fragrances and tastes for human bodies, can meet the requirements of allergic people allergic to fish products, and enriches the variety of aquatic products.
Description
Technical Field
The invention relates to the technical field of food processing, in particular to hypoallergenic cod surimi fermented by lactobacillus helveticus and a preparation method thereof.
Background
The cod is used as high-protein low-calorie fish, is a high-quality animal protein source, can provide various essential amino acids for human beings, has short muscle fiber of fish, high moisture content and tender meat, is easier to absorb than the meat of livestock, and therefore the demand of cod products is increased year by year. As one of eight major allergens, the fish can increase the allergic symptoms of the fish when the consumption is increased. The allergens responsible for the allergic symptoms of fish are diverse in variety, including the major allergen beta-parvalbumin and some minor allergens aldolase, beta-enolase, alpha-collagen, etc.
According to the prior researches, the technology for reducing and processing the aquatic product allergen can be divided into three types of physical methods, chemical methods and biological methods. The physical method mainly comprises heat treatment, irradiation treatment, pulse ultraviolet treatment, cold plasma treatment, ultrasonic treatment and high-pressure treatment, the chemical method mainly comprises Maillard reaction and enzyme method, and the biological method mainly comprises microbial fermentation and genetic engineering technology.
Fermentation is the process of converting organic matter into a product by means of biocatalysts (microbial cells or enzymes) using the metabolic activity of microorganisms. Fermentation is one of the traditional methods of food processing and preservation. The fermentation is not only helpful for digestion and absorption of food, but also can decompose food protein with antigenic property to a certain extent or denature the food protein into small molecular polypeptide and amino acid, thereby destroying certain epitope to achieve the aim of reducing sensitization, and simultaneously, the fermentation also has the advantages of improving the texture, flavor and nutritional value of the product, increasing stability and the like.
The low-sensitization products prepared by utilizing probiotics fermentation are concentrated on products such as fermented soybean products, fermented cow milk products, fermented wheat products and the like, and the low-sensitization fermented minced fillet products are mainly applied with probiotics such as lactobacillus plantarum and the like. Lactobacillus helveticus is a probiotic which exists in a large amount in the intestinal tract of human bodies, and has higher activity function of hydrolyzed protein than a plurality of probiotics, and the products related to lactobacillus helveticus on the market are basically limited to dairy products. Whether the lactobacillus helveticus fermentation can reduce the sensitization of the cod surimi allergic protein is still to be researched and has great significance.
Disclosure of Invention
The invention aims to provide a processing method for reducing the sensitization of the cod surimi allergic protein, which aims to solve the problem that whether the sensitization of the cod surimi allergic protein can be reduced by lactobacillus helveticus fermentation is mentioned in the background.
In order to achieve the above object, the present invention provides the following technical solutions:
the invention provides a preparation method of hypoallergenic cod surimi fermented by lactobacillus helveticus,
mixing the minced cod, salt, glucose and lactobacillus helveticus, and fermenting;
the lactobacillus helveticus is one or more of lactobacillus helveticus Lh187926, lactobacillus helveticus Lh191404 and lactobacillus helveticus Lh189793.
Preferably, the fermentation time is 50-70 h, and the fermentation temperature is 28-32 ℃.
Preferably, the addition amount of the salt is 1.5-2.5% of the mass of the cod surimi.
Preferably, the addition amount of the glucose is 2.5-3.5% of the mass of the cod surimi.
Preferably, the addition amount of the lactobacillus helveticus is 4-6% of the mass of the cod surimi.
Preferably, the cod is one of atlantic cod, pacific cod, holly cod and walleye cod.
Preferably, the lactobacillus helveticus Lh187926 has a bacterial content of 6.5-7.5 logcfu/g.
Preferably, the lactobacillus helveticus Lh191404 has a bacterial content of 8-10 log cfu/g.
Preferably, the lactobacillus helveticus Lh189793 has a bacterial content of 8-9 log cfu/g.
The invention also provides the hypoallergenic cod surimi prepared by the preparation method of the hypoallergenic cod surimi fermented by the lactobacillus helveticus.
Compared with the prior art, the invention has the following beneficial effects:
(1) According to the invention, the sensitization of the cod chyme allergic protein is reduced by adopting the processing modes of fermentation of different lactobacillus helveticus strains, the optimal strain is preferably obtained, the allergen reduction efficiency is improved, and a better reduction effect is obtained; the prepared hypoallergenic cod surimi is delicious in taste and rich in nutrition, can provide various essential amino acids, colors, fragrances and tastes for human bodies, can meet the requirements of allergic people allergic to fish products, enriches the variety of aquatic products, and promotes the economic development of aquatic products in China.
(2) The processing technology can effectively reduce the major allergen parvalbumin, the minor allergen aldolase, the beta-enolase and the alpha-collagen of the fish, and the obtained minced fish product has the sensibilization reduced by more than 60 percent and obvious reduction effect through the analysis of an indirect enzyme-linked immunosorbent assay.
(3) Compared with the existing allergen reduction technology, the invention is characterized in that: the biological method with milder fish quality damage is adopted, so that a better fish allergen reduction effect is realized. The selected starter has good probiotic characteristics and safety, realizes remarkable reduction effect of the fish allergen and ensures the quality of minced fillet. In addition, the fermentation system microorganism can produce a large amount of enzymes and acids, so that the gastrointestinal digestion function of the human body is enhanced, the further degradation of allergic proteins in the human body is facilitated, and the potential sensitization of fish allergic proteins is reduced. This provides technical support for the development of high quality surimi processing products.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings that are required to be used in the embodiments or the description of the prior art will be briefly described below, and it is obvious that the drawings in the following description are only embodiments of the present invention, and that other drawings can be obtained according to the provided drawings without inventive effort for a person skilled in the art.
FIG. 1 shows the variation of the total number of Lactobacillus and colony during fermentation of Lactobacillus helveticus with different strains a: lactobacillus helveticus Lh187926; b: lactobacillus helveticus Lh191404; c: lactobacillus helveticus Lh189793;
FIG. 2 shows the pH change during fermentation of different strains of Lactobacillus helveticus;
FIG. 3 comparison of SDS-PAGE of protein fractions obtained by fermentation of different strains of Lactobacillus helveticus with immunoblots a: lactobacillus helveticus Lh187926; b: lactobacillus helveticus Lh191404; c: lactobacillus helveticus Lh189793;
FIG. 4 variation of IgG binding capacity of protein fraction a obtained by fermentation of different strains of Lactobacillus helveticus: lactobacillus helveticus Lh187926; b: lactobacillus helveticus Lh191404; c: lactobacillus helveticus Lh189793.
Detailed Description
The invention provides a preparation method of hypoallergenic cod surimi fermented by lactobacillus helveticus,
mixing the minced cod, salt, glucose and lactobacillus helveticus, and fermenting;
in the invention, the lactobacillus helveticus is one or more of lactobacillus helveticus Lh187926, lactobacillus helveticus Lh191404 and lactobacillus helveticus Lh189793; preferably lactobacillus helveticus Lh187926.
In the invention, the fermentation time is 50-70 h; preferably 54 to 66 hours; further preferably 58 to 62 hours; more preferably 60h.
In the invention, the fermentation temperature is 28-32 ℃; preferably 29 to 31 ℃; further preferably 30 ℃.
In the invention, the addition amount of the salt is 1.5-2.5% of the mass of the cod surimi; preferably 1.7 to 2.3%; further preferably 1.9 to 2.1%; more preferably 2%.
In the invention, the addition amount of the glucose is 2.5-3.5% of the mass of the cod surimi; preferably 2.7 to 3.3%; more preferably 2.9 to 3.1%; more preferably 3%.
In the invention, the addition amount of the lactobacillus helveticus is 4-6% of the mass of the cod surimi; preferably 4.4 to 5.6%; further preferably 4.8 to 5.2%; more preferably 5%.
In the present invention, the cod is one of atlantic cod, pacific cod, holly cod and walleye cod; preferably atlantic cod.
In the invention, the bacterium content of the lactobacillus helveticus Lh187926 is 6.5-7.5 logcfu/g; preferably 7logcfu/g.
In the invention, the lactobacillus helveticus Lh191404 has a bacterial content of 8-10 logcfu/g, preferably 9logcfu/g.
In the invention, the lactobacillus helveticus Lh189793 has a bacterial content of 8-9 logcfu/g, preferably 8.5logcfu/g.
The invention also provides the hypoallergenic cod surimi prepared by the preparation method of the hypoallergenic cod surimi fermented by the lactobacillus helveticus.
In the invention, the lactobacillus helveticus is lactobacillus helveticus Lh187926 purchased from the north nano organism, and the lactobacillus helveticus is prepared from the following materials in batches: BNCC18726, lactobacillus helveticus Lh191404 were purchased from North Nanophyte, batch: BNCC191404; lactobacillus helveticus Lh189793 was purchased from northna, batch: BNCC189793.
The technical solutions provided by the present invention are described in detail below with reference to examples, but they should not be construed as limiting the scope of the present invention.
In the following examples, the lactobacillus helveticus was purchased from the north nano organism, batch for lactobacillus helveticus Lh 187926: BNCC18726; lactobacillus helveticus Lh191404 was purchased from northna, batch: BNCC191404; lactobacillus helveticus Lh189793 was purchased from northna, batch: BNCC189793.
In the following examples, the treatment tool includes cleaning water and the meat grinder is sterilized.
Example 1
A preparation method of hypoallergenic cod surimi fermented by lactobacillus helveticus comprises the following steps:
(1) Inoculating Lactobacillus helveticus Lh187926 into MRS liquid culture medium, culturing at 37deg.C/200 rpm for 48 hr, passaging once, culturing at 37deg.C/200 rpm for 12 hr, freeze centrifuging at 4deg.C/min for 15min, collecting thallus, washing with sterile physiological saline for 2 times, suspending in 5ml sterile physiological saline again to obtain activated bacterial suspension with bacterial content of 7log cfu/g, and preserving at 4deg.C for use.
(2) Removing head and tail of frozen Atlantic cod, removing viscera, cleaning with sterile water, and collecting middle section back muscle of cod; beating the back muscle of the middle section into minced fillet by a meat grinder;
(3) After mixing the cod surimi, 2% salt, 3% glucose, 5% lactobacillus helveticus Lh187926, fermenting at 30 ℃ for 60 hours, and after fermentation, storing the surimi sample in a refrigerator at-80 ℃ to stop the reaction.
Example 2
A preparation method of hypoallergenic cod surimi fermented by lactobacillus helveticus comprises the following steps:
(1) Inoculating Lactobacillus helveticus Lh191404 into MRS liquid culture medium, culturing at 37deg.C/200 rpm for 12 hr, passaging once, culturing at 37deg.C/200 rpm for 12 hr, freeze centrifuging at 4deg.C/min for 15min, collecting thallus, washing with sterile physiological saline for 2 times, suspending in 5ml sterile physiological saline again to obtain activated bacterial suspension with bacterial content of 9log cfu/g, and preserving at 4deg.C for use.
(2) Removing head and tail of frozen Atlantic cod, removing viscera, cleaning with sterile water, and collecting middle section back muscle of cod; beating the back muscle of the middle section into minced fillet by a meat grinder;
(3) After mixing the cod surimi, 2% salt, 3% glucose, 5% lactobacillus helveticus Lh187926, fermenting at 30 ℃ for 60 hours, and after fermentation, storing the surimi sample in a refrigerator at-80 ℃ to stop the reaction.
Example 3
A preparation method of hypoallergenic cod surimi fermented by lactobacillus helveticus comprises the following steps:
(1) Inoculating Lactobacillus helveticus Lh189793 into MRS liquid culture medium, culturing at 37 deg.C/200 rpm for 24 hr, passaging once, culturing at 37 deg.C/200 rpm for 12 hr, freeze centrifuging at 4 deg.C and 8000r/min for 15min, collecting thallus, washing with sterile physiological saline for 2 times, suspending in 5ml sterile physiological saline again to obtain activated bacterial suspension with bacterial content of 8.5log cfu/g, and preserving at 4 deg.C for use.
(2) Removing head and tail of frozen Atlantic cod, removing viscera, cleaning with sterile water, and collecting middle section back muscle of cod; beating the back muscle of the middle section into minced fillet by a meat grinder;
(3) After mixing the cod surimi, 2% salt, 3% glucose, 5% lactobacillus helveticus Lh187926, fermenting at 30 ℃ for 60 hours, and after fermentation, storing the surimi sample in a refrigerator at-80 ℃ to stop the reaction.
Example 4
A preparation method of hypoallergenic cod surimi fermented by lactobacillus helveticus comprises the following steps:
(1) Inoculating Lactobacillus helveticus Lh191404 into MRS liquid culture medium, culturing at 37deg.C/200 rpm for 12 hr, passaging once, culturing at 37deg.C/200 rpm for 12 hr, freeze centrifuging at 4deg.C/min for 15min, collecting thallus, washing with sterile physiological saline for 2 times, suspending in 5ml sterile physiological saline again to obtain activated bacterial suspension with bacterial content of 10log cfu/g, and preserving at 4deg.C for use.
(2) Removing head and tail of the frozen Pacific cod, removing viscera, cleaning with sterile water, and collecting middle back muscle of the cod; beating the back muscle of the middle section into minced fillet by a meat grinder;
(3) After mixing the cod surimi, 2.5% salt, 2.5% glucose, 6% lactobacillus helveticus, fermenting at 28deg.C for 70h, and after fermentation, storing surimi sample in-80deg.C refrigerator to stop reaction.
Example 5
A preparation method of hypoallergenic cod surimi fermented by lactobacillus helveticus comprises the following steps:
(1) Inoculating Lactobacillus helveticus Lh189793 into MRS liquid culture medium, culturing at 37 deg.C/200 rpm for 24 hr, passaging once, culturing at 37 deg.C/200 rpm for 12 hr, freeze centrifuging at 4 deg.C and 8000r/min for 15min, collecting thallus, washing with sterile physiological saline for 2 times, suspending in 5ml sterile physiological saline again to obtain activated bacterial suspension with bacterial content of 8.5log cfu/g, and preserving at 4 deg.C for use.
(2) Removing head and tail of the frozen Greenland cod, removing viscera, cleaning with sterile water, and collecting middle back muscle of cod; beating the back muscle of the middle section into minced fillet by a meat grinder;
(3) After mixing the cod surimi, 2% salt, 3% glucose and 5% lactobacillus helveticus, fermenting at 30deg.C for 60 hr, and after fermentation, storing surimi sample in-80deg.C refrigerator to stop reaction.
Example 6
A preparation method of hypoallergenic cod surimi fermented by lactobacillus helveticus comprises the following steps:
(1) Inoculating Lactobacillus helveticus Lh189793 into MRS liquid culture medium, culturing at 37 deg.C/200 rpm for 24 hr, passaging once, culturing at 37 deg.C/200 rpm for 12 hr, freeze centrifuging at 4 deg.C and 8000r/min for 15min, collecting thallus, washing with sterile physiological saline for 2 times, suspending in 5ml sterile physiological saline again to obtain activated bacterial suspension with bacterial content of 8.5log cfu/g, and preserving at 4 deg.C for use.
(2) Inoculating Lactobacillus helveticus Lh191404 into MRS liquid culture medium, culturing at 37deg.C/200 rpm for 12 hr, passaging once, culturing at 37deg.C/200 rpm for 12 hr, freeze centrifuging at 4deg.C/min for 15min, collecting thallus, washing with sterile physiological saline for 2 times, suspending in 5ml sterile physiological saline again to obtain activated bacterial suspension with bacterial content of 10log cfu/g, and preserving at 4deg.C for use.
(3) Removing head and tail of the frozen Greenland cod, removing viscera, cleaning with sterile water, and collecting middle back muscle of cod; beating the back muscle of the middle section into minced fillet by a meat grinder;
(4) Mixing cod minced fillet, 2% salt, 3% glucose, 5% Lactobacillus helveticus (the mass ratio of Lactobacillus helveticus Lh189793 and Lactobacillus helveticus Lh191404 is 1:1), fermenting at 30deg.C for 60h, and storing minced fillet sample in-80deg.C refrigerator after fermentation is completed to stop reaction.
Example 7
A preparation method of hypoallergenic cod surimi fermented by lactobacillus helveticus comprises the following steps:
(1) Inoculating Lactobacillus helveticus Lh189793 into MRS liquid culture medium, culturing at 37 deg.C/200 rpm for 24 hr, passaging once, culturing at 37 deg.C/200 rpm for 12 hr, freeze centrifuging at 4 deg.C and 8000r/min for 15min, collecting thallus, washing with sterile physiological saline for 2 times, suspending in 5ml sterile physiological saline again to obtain activated bacterial suspension with bacterial content of 8.5log cfu/g, and preserving at 4 deg.C for use.
(2) Inoculating Lactobacillus helveticus Lh191404 into MRS liquid culture medium, culturing at 37deg.C/200 rpm for 12 hr, passaging once, culturing at 37deg.C/200 rpm for 12 hr, freeze centrifuging at 4deg.C/min for 15min, collecting thallus, washing with sterile physiological saline for 2 times, suspending in 5ml sterile physiological saline again to obtain activated bacterial suspension with bacterial content of 10log cfu/g, and preserving at 4deg.C for use.
(3) Inoculating Lactobacillus helveticus Lh187926 into MRS liquid culture medium, culturing at 37deg.C/200 rpm for 48 hr, passaging once, culturing at 37deg.C/200 rpm for 12 hr, freeze centrifuging at 4deg.C/min for 15min, collecting thallus, washing with sterile physiological saline for 2 times, suspending in 5ml sterile physiological saline again to obtain activated bacterial suspension with bacterial content of 7log cfu/g, and preserving at 4deg.C for use.
(4) Removing head and tail of the frozen Greenland cod, removing viscera, cleaning with sterile water, and collecting middle back muscle of cod; beating the back muscle of the middle section into minced fillet by a meat grinder;
(5) After mixing cod surimi, 2% salt, 3% glucose, 5% lactobacillus helveticus (lactobacillus helveticus Lh189793, lactobacillus helveticus Lh191404 and lactobacillus helveticus Lh187926 in a mass ratio of 1:1:1), fermenting at 30 ℃ for 60 hours, and after fermentation, storing surimi samples in a refrigerator at-80 ℃ to stop the reaction.
Comparative example 1
The other methods were identical to example 1, except that the fermentation time was 12h.
Comparative example 2
The other methods are the same as in example 1, except that the fermentation time is 24 hours.
Comparative example 3
The other processes are identical to example 1, except that the fermentation time is 40h.
Experimental example 1
The different examples of the cod emulsion microflora were assayed for pH change and the results are shown in Table 1.
TABLE 1 preparation of hypoallergenic codfish emulsion microflora and pH Change by fermentation of different Lactobacillus helveticus
| Number of Lactobacillus helveticus | Colony count | pH | |
| Example 1 | 7.07±0.03 | 7.11±0.05 | 3.74±0.02 |
| Example 2 | 9.57±0.04 | 9.58±0.04 | 3.49±0.15 |
| Example 3 | 8.37±0.07 | 8.57±0.04 | 4.27±0.7 |
Units: CFU/g
The results show that the lactobacillus helveticus can ensure that the percentage of the lactobacillus helveticus in the total bacterial count reaches more than 95% in the fermentation time of 60 hours in the example 1, the example 2 and the example 3, which indicate that the lactobacillus helveticus always acts as a dominant strain in a microbiota in the fermentation process, wherein the lactobacillus helveticus number reaches 9.57+/-0.04 at most after the fermentation time of 60 hours in the example 2, and more nitrogen sources are consumed for growth, which is helpful for the degradation of the codfish chyme protein. Example 1, example 2, example 3 will reduce the pH more after 60 hours of fermentation than the unfermented minced fillet sample, wherein example 2 shows the best acid generating capacity, reducing the pH up to 3.49±0.15 after 60 hours of fermentation, rapid proliferation of lactobacillus helveticus accompanied by the production of large amounts of lactic acid, promoting pH reduction, lower pH helping to increase the safety of the fermented product, and also producing pleasant flavor, color and aroma.
Experimental example 2
Allergen extraction comprising the steps of:
2g of the fermented cod surimi product prepared in examples 1-3 was homogenized in 5 volumes of pre-chilled protein extraction buffer (0.1 MTris buffer; 0.5mM glycine; 1mM dithiothreitol; pH 10.5). Shaking for 12 hours in a shaking table at a constant temperature of 4 ℃, centrifuging at 9000rpm for 20min, and collecting supernatant. The supernatant was dialyzed against ultrapure water at 4℃for 24 hours. Finally, the protein liquid is put into a refrigerator with the temperature of minus 20 ℃ for standby.
Experimental example 3
SDS-PAGE and Westernblot experiments were performed as follows:
the fermented cod surimi protein samples prepared in Experimental example 2 were mixed with 5 Xelectrophoresis loading buffer at a ratio of 4:1 (v/v) and boiling for 7 minutes. The treated sample was then centrifuged at 12,000rpm for 3 minutes. Thereafter, the samples were loaded onto SDS-PAGE gels prepared using the AnyKDGPAGE kit, electrophoresed at 120V for 1.5 hours, and the gels were then stained with Coomassiebrililiant blue R-250 or used for the following Western blot detection.
The separation gel was washed with 20% ethanol before transferring the protein to PVDF membrane using iBlotTM2 gel transfer apparatus, and transfer blotting was performed after 7 minutes. After blocking with 5% skim milk and 1% Bovine Serum Albumin (BSA) (w/v) at 25 ℃ for 2 hours, incubation with rabbit antisera against PV (parvalbumin) (1:80000 diluted with PBST) and goat antisera against cod extract (1:20000 diluted with PBST) was performed in an incubator for 2 hours, followed by three washes with PBST for 10 minutes each. Subsequently, horseradish peroxidase (HRP) -labeled goat anti-rabbit IgG (1:10000 pbst dilution), HRP-labeled rabbit anti-goat IgG (1:10000 pbst dilution) were incubated for 2 hours at room temperature. Wash 3 times with PBST for 10 minutes each. The best signal was obtained from the BIORAD UniversalHoodII gel imaging system using ECL solution with multiple exposure times.
As shown in FIG. 3, the protein composition was changed after 60 hours of fermentation in examples 1, 2 and 3, the major allergen protein beta-parvalbumin of the fish in examples 2 and 3, the minor allergen aldolase, beta-enolase and alpha-collagen bands were made shallow, and the major allergen protein beta-parvalbumin of the fish in example 1 was hardly made shallow, and the minor allergen aldolase, beta-enolase and alpha-collagen bands were made shallow. Notably, examples 2 and 3 made the major and minor allergen proteins of the fish significantly shallower than example 1.
The immunoblots in fig. 3 reflect well the changes in antigenicity of the allergen proteins of example 1, example 2, example 3 during fermentation, and the results show that example 2 minimizes antigenicity of the major allergen β -parvalbumin, and that for the minor allergens aldolase, β -enolase, α -collagen, example 1, example 2 can all be eliminated after 60 hours of fermentation, but example 2 requires less time.
Experimental example 4
The samples of examples 1 to 3 prepared in experimental example 2 were taken for detection of the sensitization change by an indirect ELISA assay. The operation is as follows:
antigen coating: samples were diluted to 5. Mu.g/mL, 100. Mu.L per well was added to the ELISA plate and coated overnight at 4 ℃.
Washing: the washing was performed three times with PBST for 10min each.
Closing: 150 μl of 1% BSA per well was blocked at 37deg.C for 2 hours.
Washing: the washing was performed three times with PBST for 10min each.
Adding an antibody: mu.L of 1:400,000 diluted rabbit anti-PV serum and 1:100,000 diluted goat anti-cod extract per well were incubated at 37℃for 1h.
Washing: the washing was performed three times with PBST for 10min each.
Adding a secondary antibody: 100 μ LHRP-labeled goat anti-rabbit IgG (1:10,000 dilution) and HRP-labeled rabbit anti-goat IgG (1:10,000 dilution) were added to each well and incubated at 37℃for 1h.
Washing: the washing was performed three times with PBST for 10min each.
Color development: 100. Mu.l TMB color development solution was added to each well, and incubated at 37℃for 10min.
And (3) terminating: 50 μl of sulfuric acid stop solution was added to each well, and absorbance was rapidly assessed at 450nm and 630nm using a multiscan MK3 microplate reader, and each sample was repeated three times. OD value = OD 450 -OD 630 。
TABLE 2 preparation of hypoallergenic cod surimi Indirect ELISA results by fermentation of different Lactobacillus helveticus
| Small albumin IgG binding reduction/% | Sensitization decrease rate/% | |
| Example 1 | 2.42 | 52.76 |
| Example 2 | 16.48 | 67.47 |
| Example 3 | 18.44 | 45.83 |
The results show that the most reduced IgG binding capacity for the major allergen, mini albumin, is example 3, followed by example 2, with little reduction in example 1. From the standpoint of reduced overall sensitization, example 2 is the best subtractive capacity. Combining the results of Table 1 with experimental example 3, lactobacillus helveticus fermentation was able to reduce the sensitization of the cod chyme allergic proteins, with the best results of example 2.
Lactobacillus helveticus is one of the commonly used strains for yoghurt, helveticus and italian cheese emmentil and GranaPadano. In recent years, lactobacillus helveticus has found wide application in food products due to its safety and probiotic properties. Lactobacillus helveticus has strong proteolytic activity and is often used for strengthening yoghurt, and protein in dairy products can be converted into bioactive peptide through lactic acid production in the fermentation process. In addition, the extracellular polysaccharide produced by lactobacillus helveticus has various probiotic properties, such as antioxidant and anticancer effects. Therefore, lactobacillus helveticus can be regarded as a multifunctional probiotic, and has wide development prospect in food.
According to the invention, three different lactobacillus helveticus Lh18726, lh191404 and Lh189793 are selected to ferment the cod surimi independently, and two or three strains can be used for fermentation simultaneously, so that the influence of the fermentation strain on allergic protein sensitization is discussed.
The results show that the lactobacillus helveticus Lh191404 starter can effectively reduce the sensitization of the cod chyme allergic proteins, and the protein in the cod can be degraded due to fermentation, so that the spatial structure of sarcoplasmic proteins is changed, the binding capacity of the main allergen protein beta-parvalbumin, the secondary allergen aldolase, beta-enolase and alpha-collagen and antibodies is greatly reduced, and the structures of the allergic proteins are greatly changed in the fermentation process, so that conformational epitopes of the antibodies are destroyed. The invention preferably selects a proper starter, and can ensure the full growth of the strain by matching with proper inoculation amount, accessory addition amount ratio, fermentation temperature, time and other conditions, so that the minced fillet is fermented well, and the allergic protein sensitization of the freshwater fish is reduced.
The foregoing is merely a preferred embodiment of the present invention and it should be noted that modifications and adaptations to those skilled in the art may be made without departing from the principles of the present invention, which are intended to be comprehended within the scope of the present invention.
Claims (2)
1. A preparation method of hypoallergenic cod surimi fermented by lactobacillus helveticus is characterized in that,
mixing the minced cod, salt, glucose and lactobacillus helveticus, and fermenting;
the lactobacillus helveticus is one or more of lactobacillus helveticus Lh187926, lactobacillus helveticus Lh191404 and lactobacillus helveticus Lh189793;
the fermentation time is 50-70 h, and the fermentation temperature is 28-32 ℃;
the addition amount of the salt is 1.5-2.5% of the mass of the cod surimi;
the addition amount of the glucose is 2.5-3.5% of the mass of the cod surimi;
the addition amount of the lactobacillus helveticus is 4-6% of the mass of the cod surimi;
the cod is one of Atlantic cod, pacific cod, greenland cod and Alaska pollack;
the bacterium content of the lactobacillus helveticus Lh187926 is 6.5-7.5 log cfu/g;
the bacterium content of the lactobacillus helveticus Lh191404 is 8-10 log cfu/g;
the bacterium content of the lactobacillus helveticus Lh189793 is 8-9 log cfu/g.
2. The hypoallergenic cod emulsion produced by the method of producing a hypoallergenic cod emulsion fermented by lactobacillus helveticus according to claim 1.
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