CN111466434B - Tuna preservative and using method thereof - Google Patents

Tuna preservative and using method thereof Download PDF

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CN111466434B
CN111466434B CN202010409270.3A CN202010409270A CN111466434B CN 111466434 B CN111466434 B CN 111466434B CN 202010409270 A CN202010409270 A CN 202010409270A CN 111466434 B CN111466434 B CN 111466434B
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tuna
preservative
phloretin
cytidine
phosphate
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CN111466434A (en
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王彦波
傅玲琳
陈剑
王永正
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Zhejiang Gongshang University
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    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23B—PRESERVING, e.g. BY CANNING, MEAT, FISH, EGGS, FRUIT, VEGETABLES, EDIBLE SEEDS; CHEMICAL RIPENING OF FRUIT OR VEGETABLES; THE PRESERVED, RIPENED, OR CANNED PRODUCTS
    • A23B4/00—General methods for preserving meat, sausages, fish or fish products
    • A23B4/14—Preserving with chemicals not covered by groups A23B4/02 or A23B4/12
    • A23B4/18—Preserving with chemicals not covered by groups A23B4/02 or A23B4/12 in the form of liquids or solids
    • A23B4/20—Organic compounds; Microorganisms; Enzymes
    • 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)
  • Chemical & Material Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Microbiology (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • General Chemical & Material Sciences (AREA)
  • Wood Science & Technology (AREA)
  • Zoology (AREA)
  • Food Science & Technology (AREA)
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  • Meat, Egg Or Seafood Products (AREA)
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Abstract

The invention relates to a tuna preservative and a using method thereof, and the tuna preservative comprises cytidine-5 'phosphoric acid, 5' -adenylic acid and phloretin. The preservative can prolong the shelf life of tuna at low temperature by surface spraying, has good inhibition effect on the growth of pseudomonas fluorescens of specific spoilage bacteria of the tuna, the formation of biological membranes and the generation of volatile basic nitrogen in the storage and preservation processes, ensures the freshness of the tuna, and ensures that cytidine-5 'phosphate, 5' -adenylic acid and phloretin in the preservative are food-grade natural products and do not harm to human bodies.

Description

Tuna preservative and using method thereof
Technical Field
The invention relates to the technical field of aquatic product preservation, in particular to a tuna preservative and a using method thereof.
Background
Tuna meat is delicious, high in protein content, low in fat, rich in nutrition and red in meat quality, and in addition, the tuna meat lives in a relatively low-pollution offshore area and coastal water area throughout the year and is regarded as green, safe and pollution-free food, so that the tuna meat is recommended by the international nutritional society as one of three kinds of nutritional fishes in the world and is also called as seabed gold.
Tuna has developed vascular system and high free histidine content, is easily influenced by external environment to cause putrefaction phenomenon and autolysis phenomenon to cause fish body to become soft and generate bad smell. Among the factors causing the aquatic products to decay, the microbial factors are the most important reasons, the bacteria wrapped in the biofilm can not only cause the food decay, but also cause food poisoning due to toxins produced by the bacteria, the nutritive value of tuna can be reduced after the tuna decays, the sense organ is influenced, and toxic and harmful substances such as volatile basic nitrogen are produced to harm the health of human beings. Most researches show that specific putrefying bacteria play a main role in the putrefaction process, so that the growth of the dominant putrefaction bacteria and the formation of a bacterial film are inhibited in the low-temperature storage and preservation process of tuna, the generation of volatile basic nitrogen is reduced, the key of slowing down the putrefaction of the tuna is realized, and the important significance is realized on improving the economic value. The traditional preservative can increase the acidity of the organism and cause excessive consumption and loss of substances such as iodine, iron, calcium and the like in the human body, and excessive intake of the traditional preservative can cause gastroenteritis, harm to human body vital organs such as liver and kidney and possibly induce symptoms such as allergy.
Disclosure of Invention
The present invention is directed to solving, at least to some extent, one of the technical problems in the related art. Therefore, the invention aims to provide the tuna preservative. The preservative can effectively destroy the mycoderm form of pseudomonas fluorescens, inhibit the growth of pseudomonas fluorescens, further reduce the generation of volatile basic nitrogen and obviously improve the storage period of tuna.
To this end, in one aspect of the present invention, the present invention provides a tuna preservative comprising: cytidine-5 'phosphate, 5' -adenylate, and phloretin.
According to the tuna preservative provided by the embodiment of the invention, the practical test result shows that the preservative can prolong the shelf life of the tuna at a low temperature by surface spraying, has a good inhibiting effect on the growth of pseudomonas fluorescens, which are specific putrefying bacteria of the tuna, the formation of a biological membrane and the generation of volatile basic nitrogen in the storage and preservation processes, ensures the freshness of the tuna, and cytidine-5 'phosphate, 5' -adenylic acid and phloretin in the preservative are food-grade natural products and do not harm to human bodies.
In addition, the tuna preservative provided by the embodiment of the invention can also have the following additional technical characteristics:
according to the embodiment of the invention, the tuna preservative comprises 45-50% of cytidine-5 'phosphate, 36-40% of 5' -adenylic acid and 10-19% of phloretin by mass percent.
According to the embodiment of the invention, the tuna preservative comprises 48% of cytidine-5 'phosphate, 38% of 5' -adenylic acid and 14% of phloretin in percentage by mass.
According to an embodiment of the invention, the tuna preservative is adapted to inhibit the growth of specific spoilage bacteria of tuna.
According to an embodiment of the invention, the specific spoilage bacteria comprise pseudomonas fluorescens.
In another aspect of the present invention, the present invention provides a method for using a tuna preservative, comprising the following steps:
(1) Weighing cytidine-5 'phosphate, 5' -adenylic acid and phloretin, mixing uniformly, and adding water to prepare a mixed solution;
(2) Cleaning fresh tuna, spraying the mixed solution uniformly, vacuum packaging, and refrigerating at 4 deg.C.
According to the application method of the tuna preservative provided by the embodiment of the invention, the preservative is sprayed on the surface of the tuna, so that the mycoderm form of pseudomonas fluorescens can be effectively inhibited from growing, the generation of volatile basic nitrogen is further reduced, and the storage period of the tuna is remarkably prolonged.
According to an embodiment of the present invention, the concentration of the mixed solution is 3mg/mL.
Additional aspects and advantages of the invention will be set forth in part in the description which follows and, in part, will be obvious from the description, or may be learned by practice of the invention.
Drawings
FIG. 1 is a graph showing the effect of a tuna preservative on growth inhibition of Pseudomonas fluorescens according to an embodiment of the present invention;
FIG. 2 is a graph showing the effect of a tuna preservative on the morphology of a biofilm of Pseudomonas fluorescens according to an embodiment of the present invention;
FIG. 3 is a graph of the effect of a tuna preservative on the total number of colonies in tunas according to an embodiment of the present invention;
FIG. 4 is a graph of the effect of tuna preservative on the production of volatile basic nitrogen in tuna according to an embodiment of the present invention.
Detailed Description
The technical solution of the present invention is illustrated by specific examples below. It is to be understood that one or more method steps mentioned in the present invention do not exclude the presence of other method steps before or after the combination step or that other method steps may be inserted between the explicitly mentioned steps; it should also be understood that these examples are for illustrative purposes only and are not intended to limit the scope of the present invention. Moreover, unless otherwise indicated, the numbering of the various method steps is merely a convenient tool for identifying the various method steps, and is not intended to limit the order in which the method steps are arranged or the scope of the invention in which the invention may be practiced, and changes or modifications in the relative relationship may be made without substantially changing the technical content.
In order to better understand the above technical solutions, exemplary embodiments of the present invention are described in more detail below. While exemplary embodiments of the invention have been shown, it should be understood that the invention may be embodied in various forms and should not be construed as limited to the embodiments set forth herein. Rather, these embodiments are provided so that this disclosure will be thorough and complete, and will fully convey the scope of the invention to those skilled in the art.
The test materials adopted by the invention are all common commercial products and can be purchased in the market.
The instrument comprises the following steps: an electronic balance, a Kjeldahl azotometer and a laser confocal scanning microscope.
Reagent: cytidine-5' phosphate, food grade, available from Shanghai Arlatin Biotechnology GmbH; 5' -adenylic acid, food grade, available from Shanghai Aladdin Biotechnology GmbH; phloretin, food grade, was purchased from Shanghai Allantin Biotechnology, inc.
The invention will now be described with reference to specific examples, which are intended to be illustrative only and not to be limiting in any way.
Example 1
According to the mass percentage, 45 percent of cytidine-5 'phosphate, 36 percent of 5' -adenylic acid and 19 percent of phloretin are uniformly mixed, and water is added to prepare a mixed solution with the concentration of 3mg/mL.
Example 2
According to the mass percentage, 48 percent of cytidine-5 'phosphate, 38 percent of 5' -adenylic acid and 14 percent of phloretin are evenly mixed, and water is added to prepare a mixed solution with the concentration of 3mg/mL.
Example 3
According to the mass percentage, 50 percent of cytidine-5 'phosphate, 40 percent of 5' -adenylic acid and 10 percent of phloretin are uniformly mixed, and water is added to prepare a mixed solution with the concentration of 3mg/mL.
Example 4
According to the mass percentage, 48 percent of cytidine-5 'phosphate, 38 percent of 5' -adenylic acid and 14 percent of phloretin are uniformly mixed, and water is added to prepare a mixed solution with the concentration of 1 mg/mL.
Example 5
According to the mass percentage, 48 percent of cytidine-5 'phosphate, 38 percent of 5' -adenylic acid and 14 percent of phloretin are evenly mixed, and water is added to prepare a mixed solution with the concentration of 5 mg/mL.
Test example
1. Determining the inhibiting effect of the tuna preservative with different concentrations on pseudomonas fluorescens
The mixed solutions prepared in examples 1 to 5 were inoculated with Pseudomonas fluorescens, placed in a refrigerated cabinet at 4 ℃, and the OD at 600nm was measured every day and repeated 3 times to draw a growth curve.
As shown in FIG. 1, the mixed solutions prepared in examples 1 to 5 all inhibited the growth of Pseudomonas fluorescens, and the effect of example 2 on inhibiting Pseudomonas fluorescens was most significant. In the mixed solution prepared in example 2, pseudomonas fluorescens hardly grew for the first four days until OD was reached at day 10600It reached 0.2336, which was much lower than 1.0555 of the control group. The effect of inhibiting Pseudomonas fluorescens is most pronounced in example 2. In the following experiments for measuring the pseudomonad fluorescens membrane morphology, the total number of colonies, and volatile basic nitrogen in the mixed solution versus tuna fillets, example 2 was used as the experimental group, and the untreated group was used as the control group.
2. Effect on the morphology of Pseudomonas fluorescens biofilm
Pseudomonas fluorescens was inoculated into the mixed solution and LB broth of example 2, respectively, left to stand for 24 hours, washed 3 times with sterile ultrapure water, stained with green fluorescent nucleic acid dye Syto9, incubated at room temperature for 20min under dark conditions, followed by aspiration of the dye, and gently rinsed twice with sterile ultrapure water to remove excess dye and free cells. A series of two-dimensional parallel sections are scanned every 1 mu m by using a Z-step mode in a laser confocal scanning microscope, the thickness of a scanned image from zero to zero is the thickness of the mycoderm, and the thickness is compared with that of a control group.
The result is shown in fig. 2, the content of the biofilm produced by pseudomonas fluorescens in the control group is large, the dyed green color is deep, the thickness of the biofilm is large, the density of the biofilm is remarkably reduced after the mixed solution is added, the maximum thickness of the biofilm is also remarkably reduced, and the green color is gradually reduced. Experimental results show that the tuna preservative can obviously inhibit the formation of mycoderm of pseudomonas fluorescens.
3. Preservation experiment of tuna
Pseudomonas fluorescens is a particular putrefactive bacterium in tuna. In the low-temperature storage process, the cold-resistant and salt-resistant pseudomonas fluorescens can continuously grow to decompose fat and protein in tissues of the tuna, so that the tuna is putrefied and has reduced quality, and the formed biofilm can continuously pollute the product and shorten the shelf life of the product.
After purchasing commercially available fresh tunas, cutting the tunas into fillets, soaking the fillets in 75% alcohol for 60s, washing the fillets with sterile water for 2-3 times, then washing the fillets with sterile physiological saline with four times of volume for 2-3 times, then irradiating the fillets with ultraviolet rays in a sterile workbench for 30-60 min, and finally drying the water on the surfaces of the tunas with sterile wind to obtain the sterile fillets. Inoculating the fish fillets with a suspension of Pseudomonas fluorescens at a suitable concentration to give a final bacterial concentration of about 104CFU/g. Then the samples were distributed in sterile trays and stored in a 4 ℃ incubator with fillets inoculated with sterile normal saline as a control group. Samples were taken periodically during storage to determine the total number of colonies and volatile basic nitrogen.
Determination of the total number of colonies: in a tuna cold storage experiment with a period of 10 days, the total number of colonies is determined every 2 days, and specifically, the number of the fish fillets which are cold stored for corresponding time is determined according to the following ratio of 1:10 preparing the fish juice. The detection method adopting the viable bacteria CTC-flow cytometer comprises the following steps: taking fish juice cultured for a corresponding time, centrifuging at 12000r/min for 10min, removing supernatant, and resuspending the bacteria solution with 1mL sterile PBS or normal saline. CTC-flow cytometer total colony number measuring method: mu.L of the sample was added to a 96-well plate, 4. Mu.L of 50mM CTC was added thereto, and incubated at 37 ℃ in the dark for 3 hours, followed by counting by a flow cytometer. A200. Mu.L sample was added to a 96-well plate without CTC as a negative control. Flow cytometerThe parameters are set as follows: FL1 was 4.11, FL2 was 6.29, FL3 was 5.47, and the volume per measurement was 50. Mu.L. The optimal reaction conditions for CTC staining were: the final concentration of CTC is 2mM, the incubation is carried out for 3h at 28 ℃ in the dark, and the minimum detection limit of the method is 103CFU/g。
The result of the total number of the bacterial colonies is shown in figure 3, compared with a control group, the growth of pseudomonas fluorescens can be delayed for 4 days by adding the mixed solution, the number of the viable bacteria is reduced by 3 logs CFU/mL in a 4-degree low-temperature storage period of 4 days, the total number of the bacterial colonies is obviously influenced by the tuna preservative, and the growth of the pseudomonas fluorescens is influenced by the preservative presumably influencing the mycoderm capacity of the pseudomonas fluorescens.
Determination of volatile basic nitrogen: volatile basic nitrogen measurements were performed on tuna during storage, and the results are shown in table 1, with data being the average of 3 measurements. Method for measuring volatile basic nitrogen: mashing the fish sample by using a tissue triturator, weighing 5g of the mashed fish sample into a conical flask, adding 45mL of perchloric acid solution, homogenizing for 2min, filtering by using filter paper, and storing the liquid to be detected at 4 ℃. The detection method is carried out by referring to national standard TVB-N, and the detection is carried out once every 2 days.
As can be seen from table 1, compared with the control group, the content of volatile basic nitrogen in the fish meat of the experimental group is significantly lower, and the preservation period of the experimental group is prolonged compared with that of the control group by taking volatile basic nitrogen of less than 20 as the secondary freshness standard; similarly, fig. 4 can more intuitively illustrate that the mixed solution can significantly inhibit the generation of volatile basic nitrogen, thereby illustrating that the tuna preservative of the present application has a good preservation effect.
TABLE 1 volatile basic nitrogen content (mg/100 g) in fish meat
Figure BDA0002492582100000051
To sum up, the tuna preservative can effectively reduce the total number of bacterial colonies in the tuna during storage, further reduce the generation of volatile basic nitrogen, slow down the deterioration and improve the product quality.
In the description of the specification, reference to the description of "one embodiment," "some embodiments," "an example," "a specific example," or "some examples" or the like means that a particular feature, structure, material, or characteristic described in connection with the embodiment or example is included in at least one embodiment or example of the invention. In this specification, the schematic representations of the terms used above should not be understood to necessarily refer to the same embodiment or example. Furthermore, the particular features, structures, materials, or characteristics described may be combined in any suitable manner in any one or more embodiments or examples. Furthermore, various embodiments or examples described in this specification can be combined and combined by those skilled in the art.
Although embodiments of the present invention have been shown and described above, it is understood that the above embodiments are exemplary and should not be construed as limiting the present invention, and that variations, modifications, substitutions and alterations can be made to the above embodiments by those of ordinary skill in the art within the scope of the present invention.

Claims (7)

1. A tuna preservative is characterized by comprising: cytidine-5 'phosphate, 5' -adenylate, and phloretin.
2. The tuna preservative according to claim 1, comprising 45 to 50% by mass of cytidine-5 'phosphate, 36 to 40% by mass of 5' -adenylic acid, and 10 to 19% by mass of phloretin.
3. The tuna preservative according to claim 2, comprising 48% cytidine-5 'phosphate, 38% 5' -adenylate and 14% phloretin by mass.
4. The tuna preservative according to any one of claims 1 to 3, which is suitable for inhibiting the growth of specific spoilage bacteria of tuna.
5. The tuna preservative of claim 4 wherein the specific spoilage bacteria comprise pseudomonas fluorescens.
6. A method of using the tuna preservative according to any one of claims 1 to 5, comprising the steps of:
(1) Weighing cytidine-5 'phosphate, 5' -adenylic acid and phloretin, mixing uniformly, and adding water to prepare a mixed solution;
(2) Cleaning fresh tuna, spraying the mixed solution uniformly, vacuum packaging, and refrigerating at 4 deg.C.
7. The method for using the tuna preservative according to claim 6, wherein the concentration of the mixed solution is 3mg/mL.
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Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
根皮素对冷藏三文鱼的保鲜效果研究;房杰;《中国优秀博硕士学位论文全文数据库（硕士）工程科技I辑》;20170515;第34-35页4.3.1样品处理 *

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