CN106962452B - Freshwater fish flesh preservative and preservation method - Google Patents
Freshwater fish flesh preservative and preservation method Download PDFInfo
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- CN106962452B CN106962452B CN201710221522.8A CN201710221522A CN106962452B CN 106962452 B CN106962452 B CN 106962452B CN 201710221522 A CN201710221522 A CN 201710221522A CN 106962452 B CN106962452 B CN 106962452B
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Classifications
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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
- A23B4/10—Coating with a protective layer; Compositions or apparatus therefor
-
- 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
- A23L3/00—Preservation of foods or foodstuffs, in general, e.g. pasteurising, sterilising, specially adapted for foods or foodstuffs
- A23L3/34—Preservation of foods or foodstuffs, in general, e.g. pasteurising, sterilising, specially adapted for foods or foodstuffs by treatment with chemicals
- A23L3/3454—Preservation of foods or foodstuffs, in general, e.g. pasteurising, sterilising, specially adapted for foods or foodstuffs by treatment with chemicals in the form of liquids or solids
- A23L3/3463—Organic compounds; Microorganisms; Enzymes
- A23L3/3472—Compounds of undetermined constitution obtained from animals or plants
-
- 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)
- Food Science & Technology (AREA)
- Polymers & Plastics (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Health & Medical Sciences (AREA)
- Nutrition Science (AREA)
- Microbiology (AREA)
- Botany (AREA)
- Wood Science & Technology (AREA)
- Zoology (AREA)
- Agricultural Chemicals And Associated Chemicals (AREA)
- Meat, Egg Or Seafood Products (AREA)
Abstract
A freshwater fish flesh preservative and a preservation method are characterized in that: the red yeast rice wine after fermentation for 15 days at the constant temperature of 25 ℃ comprises an extracting solution which is prepared by mixing dark plum, nutmeg, clove and garlic in equal volume and extracting the red yeast rice wine, wherein the material-liquid ratio of the dark plum extracting solution is 67-72 g/L, the material-liquid ratio of the nutmeg extracting solution is 90-100 g/L, the material-liquid ratio of the clove extracting solution is 40-50 g/L, and the material-liquid ratio of the garlic extracting solution is 125-143 g/L. The components of the preservative prepared by the invention have positive synergy to enhance the bacteriostatic effect. The preservative disclosed by the invention can be used for enabling the fish preservation effect to be optimal and effectively prolonging the shelf life of the fish for 5-6 days. The red yeast rice wine is positively cooperated with the spice component to enhance the antibacterial effect, and simultaneously can ensure good flavor and excellent fish meat quality and sense. The red yeast rice wine is health care wine, and the natural spice is also a food component, so the preservative has high efficiency, nutrition, health, no side effect and good development and application prospects.
Description
Technical Field
The invention belongs to the technical field of aquatic product preservation, and particularly relates to a freshwater fish flesh preservative and a fresh-keeping method.
Background
The freshwater fish resources in China are rich, and the method is the country with the largest aquatic product yield in the world. The freshwater fish not only has tender meat quality, but also has rich nutrition, and is always well received by consumers. However, the market value is very limited due to the fact that freshwater fish cannot be preserved, the freshwater fish industry is limited by storage and processing conditions, the putrefaction rate is over 30% every year, the processing rate is less than 10%, and great loss is caused to fishermen and fish product processing enterprises.
At present, the fish meat has the following processing, preservation and sale channels: a small number of the live-keeping ships are adopted to transport the live-keeping ships to destinations for sale; the fresh-keeping method is characterized in that fresh-keeping preservation is partially adopted, namely, a layer of crushed ice and a layer of fish are preserved and sealed in a whole box and are packaged, the fresh-keeping fish in the mode generally mainly enters supermarkets and vegetable yards, the freshness can be maintained for 3-4 days in the winter transportation, if the foam box is placed into a refrigeration house for refrigeration at 0-4 ℃, the freshness index after 13-14 days can still reach the first grade, but the quality is difficult to guarantee after the period is exceeded; the caught fish is conveyed to a processing factory, is processed into a frozen product after being plated with ice coating, is frozen in a refrigeration house and is sold by machine; the salt content of the dried product is generally high, but the fresh and tender texture of the fresh fish muscle is lacked, and the requirements of modern diet consumption on nature, greenness and health are not met. The processing and preservation can not guarantee the freshness, flavor and texture of the fresh fish for a long time.
Disclosure of Invention
This section is for the purpose of summarizing some aspects of embodiments of the invention and to briefly introduce some preferred embodiments. In this section, as well as in the abstract and the title of the invention of this application, simplifications or omissions may be made to avoid obscuring the purpose of the section, the abstract and the title, and such simplifications or omissions are not intended to limit the scope of the invention.
The invention is provided in view of the above and/or the technical blank of the existing fresh-water fish flesh preservation.
Therefore, one of the purposes of the invention is to solve the defects in the prior art and provide a healthy and efficient fresh-water fish flesh preservative.
In order to solve the technical problems, the invention provides the following technical scheme: the red yeast rice wine after fermentation for 15 days at the constant temperature of 25 ℃ comprises an extracting solution which is prepared by mixing dark plum, nutmeg, clove and garlic in equal volume and extracting the red yeast rice wine, wherein the material-liquid ratio of the dark plum extracting solution is 67-72 g/L, the material-liquid ratio of the nutmeg extracting solution is 90-100 g/L, the material-liquid ratio of the clove extracting solution is 40-50 g/L, and the material-liquid ratio of the garlic extracting solution is 125-143 g/L.
As a preferred scheme of the freshwater fish flesh preservative provided by the invention, the freshwater fish flesh preservative comprises the following components in parts by weight: the preparation method of the red yeast rice wine comprises the following steps of soaking: soaking the sticky rice in clear water for 3.5-4.5 hours; and (3) cooking: after being easily crushed by hand, the glutinous rice is put into a pot for cooking, and when the glutinous rice is cooked to be eighty percent cooked, the glutinous rice is taken out and cooled; mixing: uniformly mixing red yeast rice and glutinous rice according to the ratio of 1: 5-1: 6, putting the mixture into an aseptic jar, and adding water, wherein the ratio of water to rice is 1.5: 1-2: 1; fermentation: placing the jar in a thermostat at 25 deg.C, and fermenting for 15 days to obtain red rice wine.
As a preferred scheme of the freshwater fish flesh preservative provided by the invention, the freshwater fish flesh preservative comprises the following components in parts by weight: the dark plum extract is characterized in that the material-to-liquid ratio of total organic acid is 23-25 g/L.
As a preferred scheme of the freshwater fish flesh preservative provided by the invention, the freshwater fish flesh preservative comprises the following components in parts by weight: the nutmeg extract is prepared from nutmeg volatile oil, wherein the material-liquid ratio of nutmeg volatile oil is 26-29 g/L.
As a preferred scheme of the freshwater fish flesh preservative provided by the invention, the freshwater fish flesh preservative comprises the following components in parts by weight: the clove extracting solution is prepared from 3.6-4.5 g/L eugenol; the garlic extract is prepared from 0.36-0.42 g/L allicin.
The invention also aims to provide a fresh-keeping method of freshwater fish flesh, which is simple and feasible and is suitable for industrial production and popularization.
In order to solve the technical problems, the invention provides the following technical scheme: a fresh-keeping method for freshwater fish flesh is characterized by comprising the following steps: the method comprises the steps of killing freshwater fish, washing the killed freshwater fish with water, slicing, rinsing, draining, soaking the freshwater fish meat in the freshwater fish meat preservative of any one of claims 1-5, packaging and refrigerating, and specifically comprises the steps of killing and washing the fresh freshwater fish, and cutting the fresh freshwater fish into fish slices with the thickness of 1-2 cm and smooth surfaces; washing the fillets in precooled sterile water for 10-15 s, and draining at the temperature of 0-4 ℃; soaking the drained fillets in a pre-cooled coating preservative for 30-60 min, wherein the volume ratio of the fillets to the freshwater fish preservative is about 1: 3; and (4) sealing, packaging and refrigerating the fillets soaked with the freshwater fish flesh preservative.
As a preferable scheme of the fresh-keeping method of the freshwater fish meat, the method comprises the following steps: the dark plum extracting solution is extracted from dark plum, and the extraction method of the dark plum extracting solution comprises the steps of crushing and sieving dark plum, dissolving the crushed dark plum in red kojic rice wine according to the liquid-material ratio of 14-15 ml/g, leaching for 70-90 min by using an ultrasonic cleaner with the temperature of 55 ℃, the frequency of 40kHz and 200W, and filtering to obtain supernatant, namely the dark plum extracting solution.
As a preferable scheme of the fresh-keeping method of the freshwater fish meat, the method comprises the following steps: the nutmeg extract is extracted from nutmeg, and the extraction method of the nutmeg extract comprises the steps of crushing and sieving nutmeg, dissolving the nutmeg in red kojic rice wine according to the liquid-material ratio of 10-11 ml/g, extracting for 2-2.5 min by using an ultrasonic microwave instrument with the frequency of 40kHz and 250-300W, and filtering to obtain supernatant, namely the nutmeg extract.
As a preferable scheme of the fresh-keeping method of the freshwater fish meat, the method comprises the following steps: the clove extracting solution is extracted from clove, and the extracting method of the clove extracting solution comprises the steps of crushing and sieving clove, dissolving the crushed clove in red kojic rice wine according to the liquid-material ratio of 20-25 ml/g, leaching for 60-90 min by using an ultrasonic cleaner with the temperature of 55-65 ℃, the frequency of 40kHz and 200W, and filtering to obtain supernatant, namely the clove extracting solution.
As a preferable scheme of the fresh-keeping method of the freshwater fish meat, the method comprises the following steps: the extraction method of the garlic extract comprises the steps of crushing and sieving garlic, dissolving the crushed garlic in red kojic rice wine according to the liquid-material ratio of 7-8 ml/g, leaching for 20-30 min by using an ultrasonic cleaner with the temperature of 30-40 ℃, the frequency of 40kHz and the power of 48-72W, and filtering to obtain supernatant, namely the garlic extract.
The invention has the following beneficial effects:
(1) the components of the preservative prepared by the invention have positive synergy to enhance the bacteriostatic effect. The preservative disclosed by the invention can be used for enabling the fish preservation effect to be optimal and effectively prolonging the shelf life of the fish for 5-6 days.
(2) The red yeast rice wine is positively cooperated with the spice component to enhance the antibacterial effect, and simultaneously can ensure good flavor and excellent fish meat quality and sense.
(3) The red yeast rice wine is health care wine, and the natural spice is also a food component, so the preservative has high efficiency, nutrition, health, no side effect and good development and application prospects.
Detailed Description
In order to make the aforementioned objects, features and advantages of the present invention comprehensible, embodiments accompanied with examples are described in detail below.
In the following description, numerous specific details are set forth in order to provide a thorough understanding of the present invention, but the present invention may be practiced in other ways than those specifically described and will be readily apparent to those of ordinary skill in the art without departing from the spirit of the present invention, and therefore the present invention is not limited to the specific embodiments disclosed below.
Furthermore, reference herein to "one embodiment" or "an embodiment" means that a particular feature, structure, or characteristic described in connection with the embodiment is included in at least one implementation of the invention. The appearances of the phrase "in one embodiment" in various places in the specification are not necessarily all referring to the same embodiment, nor are separate or alternative embodiments mutually exclusive of other embodiments.
Example 1
Fresh-keeping agent for freshwater fish flesh
Preparing red yeast rice wine:
soaking: soaking the sticky rice in clear water for 3.5-4.5 hours;
and (3) cooking: after being easily crushed by hand, the glutinous rice is put into a pot for cooking, and when the glutinous rice is cooked to be eighty percent cooked, the glutinous rice is taken out and cooled;
mixing: uniformly mixing red yeast rice and glutinous rice according to the ratio of 1: 5-1: 6, putting the mixture into an aseptic jar, and adding water, wherein the ratio of water to rice is 1.5: 1;
fermentation: placing the jar in a thermostat at 25 deg.C, and fermenting for 15 days to obtain red rice wine.
Preparing a spice extracting solution:
preparing a dark plum extracting solution: pulverizing mume fructus, sieving, dissolving in red rice wine at a liquid-material ratio of 14ml/g, extracting with ultrasonic cleaner at 55 deg.C and frequency of 40kHz and 200W for 80min, filtering, and collecting supernatant to obtain mume fructus extract.
Preparation of a nutmeg extract: pulverizing semen Myristicae, sieving, dissolving in red rice wine at a liquid-material ratio of 10ml/g, extracting with ultrasonic microwave instrument at 40kHz and 250W for 2.5min, filtering, and collecting supernatant to obtain semen Myristicae extract.
Preparing a clove extracting solution: pulverizing flos Caryophylli, sieving, dissolving in red rice wine at a liquid-to-material ratio of 20ml/g, extracting with ultrasonic cleaner at 60 deg.C and frequency of 40kHz and 200W for 80min, filtering, and collecting supernatant to obtain flos Caryophylli extractive solution.
Preparing garlic extract: pulverizing Bulbus Allii, sieving, dissolving in red rice wine at a liquid-to-material ratio of 7ml/g, extracting with ultrasonic cleaner at 35 deg.C and frequency of 40kHz and 50W for 25min, filtering, and collecting supernatant to obtain Bulbus Allii extract.
Preparing a preservative:
and mixing the prepared dark plum, nutmeg, clove and garlic extract in equal volume to prepare the freshwater fish flesh preservative of the invention.
Fresh-keeping of fish meat by using fresh-keeping agent
Slaughtering freshwater fish, washing with water, slicing, rinsing, draining, soaking with the prepared freshwater fish fresh-keeping agent, packaging and refrigerating,
killing and cleaning fresh freshwater fish, and cutting the fish into fish slices with the thickness of 1-2 cm and smooth surfaces; washing the fillets in precooled sterile water for 10-15 s, and draining at the temperature of 0-4 ℃; soaking the drained fish fillets in a pre-cooled coating preservative for 45min, wherein the volume ratio of the fish fillets to the fresh-water fish preservative is about 1: 3;
and (4) sealing, packaging and refrigerating the fillets soaked with the freshwater fish flesh preservative.
Example 2
And 4, determining the extraction rate of dark plum, nutmeg, clove and garlic extract:
and (3) determining the extraction rate of the dark plum extract:
preparation of dark plum sample 1: pulverizing mume fructus, sieving, dissolving in red rice wine at a liquid-material ratio of 14ml/g, extracting with ultrasonic cleaner at 55 deg.C and frequency of 40kHz and 200W for 70min, filtering, and collecting supernatant to obtain mume fructus extract.
Preparation of dark plum sample 2: pulverizing mume fructus, sieving, dissolving in red rice wine at a liquid-material ratio of 15ml/g, extracting with ultrasonic cleaner at 55 deg.C and frequency of 40kHz and 200W for 80min, filtering, and collecting supernatant to obtain mume fructus extract.
Preparation of dark plum sample 3: pulverizing mume fructus, sieving, weighing, dissolving in red rice wine at a liquid-material ratio of 15ml/g, extracting with ultrasonic cleaner at 55 deg.C and frequency of 40kHz and 200W for 90min, filtering, and collecting supernatant to obtain mume fructus extract.
Determining the extraction rate of the dark plum sample: precisely measuring 1ml of each sample extracting solution, placing the sample extracting solution in an erlenmeyer flask, adding 10ml of water, adding 2 drops of phenolphthalein indicator solution, and titrating the solution by using NaOH titration solution (0.1mol/L) until the solution is changed from colorless to red.
The extraction rate of the effective components is determined according to the following formula:
extraction rate ═ CVNaOH×Mr/m×3000]×100％
Wherein the content of the first and second substances,
c is the solution concentration of NaOH, and the unit is mol/L;
v is the volume of NaOH solution consumed, and the unit is L;
Mrthe molar mass of the organic acid is 192 g/mol;
m is the mass of the dark plum and the unit is g.
The results of the measurements are given in table 2.1 below:
TABLE 2.1 results of extraction of dark plum
And (3) measuring the extraction rate of the myristica fragrans extract:
preparation of nutmeg sample 1: pulverizing semen Myristicae, sieving, weighing, dissolving in red rice wine at a liquid-material ratio of 10ml/g, extracting with ultrasonic microwave instrument at 40kHz and 250W for 2.5min, filtering, and collecting supernatant to obtain semen Myristicae extract.
Preparation of nutmeg sample 2: pulverizing semen Myristicae, sieving, weighing, dissolving in red rice wine at a liquid-material ratio of 11ml/g, extracting with 40kHz 250W ultrasonic microwave instrument for 2min, filtering, and collecting supernatant to obtain semen Myristicae extract.
Preparation of nutmeg sample 3: pulverizing semen Myristicae, sieving, weighing, dissolving in red rice wine at a liquid-material ratio of 11ml/g, extracting with 40kHz 300W ultrasonic microwave instrument for 2min, filtering, and collecting supernatant to obtain semen Myristicae extract.
Determination of extraction rate of nutmeg sample: concentrating the extractive solution in rotary evaporator until no red rice wine is separated out, extracting the concentrated solution with n-hexane, and rotary concentrating the extractive solution until n-hexane is completely evaporated to obtain semen Myristicae volatile oil.
The extraction rate of the nutmeg volatile oil is calculated according to the following formula:
the extraction rate of nutmeg volatile oil (volatile oil mass/raw material mass) is × 100%
The results of the measurements are given in table 2.2 below:
TABLE 2.2 myristica fragrans extraction yield results
And (3) determining the extraction rate of the clove extracting solution:
preparation of clove sample 1: pulverizing flos Caryophylli, sieving, dissolving in red rice wine at a liquid-to-material ratio of 20ml/g, extracting with ultrasonic cleaner at 55 deg.C and frequency of 40kHz and 200W for 60min, filtering, and collecting supernatant to obtain flos Caryophylli extractive solution.
Preparation of clove sample 2: pulverizing flos Caryophylli, sieving, dissolving in red rice wine at a liquid-to-material ratio of 23ml/g, extracting with ultrasonic cleaner at 60 deg.C and frequency of 40kHz and 200W for 70min, filtering, and collecting supernatant to obtain flos Caryophylli extractive solution.
Preparation of clove sample 3: pulverizing flos Caryophylli, sieving, dissolving in red rice wine at a liquid-to-material ratio of 25ml/g, extracting with ultrasonic cleaner at 65 deg.C and frequency of 40kHz and 200W for 90min, filtering, and collecting supernatant to obtain flos Caryophylli extractive solution.
And (3) determining the extraction rate of the clove sample: concentrating the extractive solution in rotary evaporator until no red rice wine is separated out, extracting the concentrated solution with n-hexane, and rotary concentrating the extractive solution until n-hexane is completely evaporated to obtain semen Myristicae volatile oil.
The extraction rate of the clove volatile oil is calculated according to the following formula:
the extraction rate of the clove volatile oil (the mass of the volatile oil/the mass of the raw materials) is × 100 percent
The results of the measurements are given in table 2.3 below:
TABLE 2.3 clove extraction yield results
And (3) determining the extraction rate of the garlic extract:
preparation of garlic sample 1: pulverizing Bulbus Allii, sieving, dissolving in red rice wine at a liquid-to-material ratio of 7ml/g, extracting with 30 deg.C ultrasonic cleaner at 40kHz and 45W for 25min, filtering, and collecting supernatant to obtain Bulbus Allii extract. Preparation of garlic sample 2: pulverizing Bulbus Allii, sieving, dissolving in red rice wine at a liquid-to-material ratio of 8ml/g, extracting with ultrasonic cleaner at 35 deg.C and frequency of 40kHz and 60W for 25min, filtering, and collecting supernatant to obtain Bulbus Allii extract.
Preparation of garlic sample 3: pulverizing Bulbus Allii, sieving, dissolving in red rice wine at a liquid-to-material ratio of 7ml/g, extracting with ultrasonic cleaner at 40 deg.C and frequency of 40kHz and 75W for 25min, filtering, and collecting supernatant to obtain Bulbus Allii extract.
Determining the extraction rate of the garlic sample: the colorimetric method is adopted for determination, and the specific operation is as follows,
taking 0.5mL of 1.0mM cysteine solution, adding 2.0mL of 2.0mM DTNB solution, diluting to 6.0mL with 50mM Hepes buffer solution, incubating at 26 deg.C for 30min, and measuring absorbance at 412nm (A)0)。
Diluting Bulbus Allii solution 50 times with 50mM Hepes buffer solution, adding 1.0mM cysteine solution 0.5mL, adding 0.5mL Bulbus Allii diluent, incubating at 26 deg.C for 10min, adding 2.0mM DTNB solution 2.0, diluting to 6.0 with 50Hepes buffer solution, incubating at 26 deg.C for 30min, and measuring absorbance (A) at 412 nm.
The calculation formula of the allicin content is as follows:
C＝△A412×d×162.26/(2×14150)；
△A412＝A0-A；
mallicin＝C×V；
Wherein C is the concentration of allicin in the garlic extract, and the unit is g/ml; v is the total volume of the garlic extract sample; d is the total dilution multiple; 162.26 is the molecular weight of allicin; 14150 is the molar extinction coefficient of the reaction product NTB (2-nitro-5-thiobenzoic acid) of DTNB and allicin at 412nm and 1cm light path.
The calculation formula of the allicin extraction rate is as follows:
garlicin extraction rate (m ═ m)Allicin/MQuality of garlic raw material)×100％
The specific results are shown in the following table 2.4:
TABLE 2.4 allicin extraction yield results
And (3) measuring the effective component liquid-liquid ratio and the raw material liquid-liquid ratio:
from the above-described extraction rate measurement experiments, it was found that the process parameters provided by the present invention are relatively stable in the extraction rate of each active ingredient. The inventor researches and discovers that the material-liquid ratio of the effective components in the product, namely the effective concentration, can be effectively controlled by controlling the extraction rate and the liquid-material ratio of the raw materials relative to the red yeast rice wine.
The specific mathematical calculations are as follows.
Setting:
the extraction rate of the effective components is a (%),
the liquid-material ratio of the raw materials to the red yeast rice wine (solvent of the extracting solution) is b (mL/g),
adding m g total raw materials, wherein the ratio of effective components to extractive solution is x (g/L),
the feed-liquid ratio of the raw material to the extract was y (g/L).
Then the following are:
the mass of the effective component is m × a (g);
the volume of the extractive solution is m × b (ml);
therefore, the ratio of the effective component to the extract solution is the mass of the effective component/the volume of the extract solution, that is,
according to the formula, the effective component feed liquid ratio is calculated as the following table 2.5:
TABLE 2.5 summary of the active ingredients
According to the data, in the preparation of the extracting solution, according to the method provided by the invention, the liquid-material ratio of the raw materials relative to the red yeast rice wine is calculated; the extraction rate of the effective components is controlled in a narrow range by controlling specific parameters of the preparation process; the control of two key parameters is integrated, so that the feed-liquid ratio of the effective components relative to the extracting solution is controlled within a target range.
Example 3
Saprophytic bacteria screening, identifying and preservative bacteriostatic experiment
Fresh carps are stunned with hard substances, and then scales, viscera and fins are removed. Washing with distilled water, washing with sterile distilled water under sterile condition for three times, and wiping fish body with 75% alcohol. After the alcohol is completely volatilized, the carp is cut by using a sterile medical scalpel, fish meat without pollution of the fish body is selected, the fish meat is cut into fish blocks of about 2 x 2cm, and each piece of sterile fish meat is about 10g and is stored in a thermostat at the temperature of 4 ℃ in a container. After the fish is refrigerated for 8 days and putrefaction of the fish occurs, taking out fish blocks, performing aseptic operation, cutting the fish with scissors, accurately weighing 25g of the fish, placing the fish into an aseptic homogenizing bag, adding 225ml of aseptic normal saline into the aseptic homogenizing bag for homogenization, taking 1ml of bacterial suspension for gradient dilution, then selecting 3 appropriate dilution gradient samples, respectively taking 1ml of the dilution gradient samples, injecting the dilution gradient samples into a culture dish, pouring the dilution gradient samples into a nutrient agar culture medium at 45 ℃ for uniform mixing, performing three parallel dilution gradients, and culturing at 37 ℃ for 48 hours.
Selecting a flat plate with more uniform bacterial colonies, selecting strains with more quantity and different bacterial colony forms in the flat plate, carrying out microscopic examination on the strains with similar bacterial colony forms, and finding out the strains with different cell forms through the microscopic examination. Meanwhile, carrying out plate streaking separation on the separated strains, and repeatedly carrying out 3-4 times to obtain purified single colonies. The single colonies obtained were numbered, a part of which was preserved on the slant and a part of which was used for the detection of the decay-causing ability.
The decay-causing capacity is measured by adopting a back grafting method, the separated 10 strains are picked by an inoculating loop under an aseptic environment, single colonies are smeared on the surface of an aseptic carp block and the inside of fish meat, the aseptic carp block and the fish meat are refrigerated in a constant-temperature incubator at 4 ℃ in an aseptic sampling bag, sensory evaluation is carried out on the aseptic carp block and the fish meat in 0, 2, 4, 6 and 8 days respectively, and whether each strain has the decay capacity is preliminarily screened.
Suspected putrefying bacteria can be obtained through sensory evaluation of the first-time back-grafting fish, then single bacterial colonies are picked by an inoculating loop under the aseptic environment and are smeared on the surface of an aseptic fish block and the inside of the fish, the aseptic fish block and the inside of the fish are placed in an aseptic sampling bag and are placed in a constant-temperature incubator at 4 ℃ for refrigeration, and the dominant putrefying bacteria are determined by measuring the content of volatile basic nitrogen of the fish in days 1, 3, 5, 7 and 9.
The method for measuring the volatile basic nitrogen refers to the standard SC/T3032-2007 determination of the volatile basic nitrogen in the aquatic products of the people's republic of China.
And then, performing morphological observation, gram staining, biochemical identification and molecular identification on the rotting strains obtained by the two-time screening respectively.
Extracting bacterial DNA, referring to a method for extracting bacterial DNA by Von Guangda and the like, adopting bacterial universal primers: -5 'AGAGAGTTT-GATC (C/A) TGGCTAG-3'; 5 '-GGTTACCTT-GTTACGACTT-3'.
The PCR amplification system was a 25. mu.l system: 9.5. mu.l of ddH2O, 0.5. mu.l of each primer pair, PCRmigx12.5. mu.l, and 2. mu.l of DNA template.
Amplification conditions: pre-denaturation at 95 ℃ for 3min, denaturation at 95 ℃ for 30s, annealing at 54 ℃ for 30s, extension at 72 ℃ for 45s, 40 cycles, extension at 72 ℃ for 7min, and preservation at 4 ℃.
Carrying out agarose gel electrophoresis on the obtained PCR amplification product, detecting, sending the amplification product meeting the requirements to a biological engineering (Shanghai) corporation, sequencing by Beijing sequencing department, carrying out comparison analysis on the obtained DNA sequence in a nucleic acid database in an NCBI website, determining the strain type according to the similarity, combining morphological characteristics and gram staining results, further identifying the strain type by a biochemical experiment, wherein the identification result is as follows: the strain 3 is Bacillus cereus, 5 is Bacillus licheniformis, 7 is Aeromonas hydrophila, and 8 is Aeromonas veroni.
The components with the same liquid-material ratio in example 1 are crushed and sieved, dissolved by using distilled water as a solvent to prepare dark plum, nutmeg, clove and garlic solutions, and mixed in equal volumes to prepare a blank mixed sample.
The preservative described in example 1 was added with red koji wine as a comparative sample 1, and a bacteriostatic circle effect test was performed by a filter paper method.
The specific experimental method comprises the following steps:
filter paper sheet method. Processing quantitative filter paper into 0.600cm round filter paper sheet with a puncher, drying and sterilizing at 165 deg.C for 2.5h, soaking in each extractive solution for half an hour, and air drying in sterile culture dish. Sucking 0.2ml of liquid culture of each dominant putrefying bacterium by using a pipette gun, uniformly coating on a nutrient agar plate, sticking filter paper sheets at proper positions of the plates by using tweezers after the bacterium liquid permeates into a culture medium and no movable water drops exist on the surface of the culture medium, sticking 6 filter paper sheets in each plate at equal intervals, performing a control test by using red kojic rice wine and sterile physiological saline, and repeating 3 times on each plate. The plate is placed in a constant temperature incubator at 37 ℃ for culturing for 18-24 hours, then the diameter of each inhibition zone is measured by a vernier caliper (micrometer), and data is recorded.
The results of the experiment are given in table 3.1 below:
TABLE 3.1 bacteriostatic test results Table
Therefore, compared with a spice blank mixed solution and a single red yeast rice wine, the preservative provided by the invention can obviously enhance the bacteriostatic effect, which shows that the spice and the red yeast rice wine have a synergistic effect on the inhibition of four dominant spoilage bacteria after being mixed.
Example 4
Fresh carps are stunned with hard substances, and then scales, viscera and fins are removed. Washing with distilled water, washing with sterile distilled water under sterile condition for three times, and wiping fish body with 75% alcohol. After the alcohol is completely volatilized, the carp is cut by using a sterile medical scalpel, fish meat without pollution of the fish body is selected and cut into fish blocks of about 2 x 2cm, and each piece of sterile fish meat is about 10 g.
The fish blocks are respectively put in the antistaling agent in the embodiment 1, the red yeast rice wine in the embodiment 3 and the blank mixed solution. Draining fish, sealing in a sterile sampling bag, and refrigerating at 4 deg.C for determination.
Selecting 6 food-professional classmates to evaluate the smell, the juice loss, the fish elasticity and the like of the fish, scoring the three indexes by 5, 3, 2 and 1 according to the standard, summing up and averaging, wherein the specific scoring standard is as the following table 4.1:
TABLE 4.1 sensory evaluation criteria for preservation experiments
Determination of the Total number of colonies
Measured according to the national standard GB 4789.2-2010
(3) Determination of volatile basic nitrogen
By using a Kjeldahl nitrogen-determination distillation device, the method refers to the standard SC/T3032 of the aquatic product industry of the people's republic of China and 2007 determination of volatile basic nitrogen in aquatic products.
The results of sensory evaluation scores of the preservation test are shown in the following table 4.2:
TABLE 4.2 sensory evaluation results of the preservation experiment
| Day 1 | Day 3 | Day 5 | Day 7 | Day 9 | Day 12 | |
| Red yeast rice wine | 15±0.00 | 13.32±0.22 | 12.90±0.37 | 10.51±0.08 | 8.93±0.12 | 4.56±0.17 |
| Blank mixing | 15±0.00 | 10.6±0.26 | 8.20±0.17 | 3.95±0.25 | 2.70±0.17 | 1.21±0.13 |
| Fresh-keeping agent | 15±0.00 | 14.89±0.41 | 14.52±0.21 | 12.85±0.12 | 10.37±0.05 | 9.28±0.07 |
The TVB-N content results are shown in Table 4.3 below:
TABLE 4.3 Table for TVB-N content (mg/100g) of preserved carp meat
| Day 1 | Day 3 | Day 5 | Day 7 | Day 9 | Day 12 | |
| Red yeast rice wine | 9.37±0.01 | 13.48±0.02 | 15.95±0.1 | 19.09±0.04 | 23.79±0.02 | 26.80±0.03 |
| Blank mixing | 9.51±0.08 | 19.07±0.04 | 26.14±0.5 | 32.761±0.02 | 41.91±0.19 | 52.1±0.15 |
| Fresh-keeping agent | 9.30±0.02 | 9.69±0.02 | 10.03±0.01 | 12.28±0.018 | 15.05±0.06 | 18.87±0.32 |
Note: the TVB-N content of the preserved carp flesh is mg/100g
The analysis results of the total number of colonies of the preserved carp flesh are shown in the following table 4.4:
TABLE 4.4 analysis Table of the total number of colonies of carp flesh
| Day 1 | Day 3 | Day 5 | Day 7 | Day 9 | Day 12 | |
| Red yeast rice wine | 3.45×103 | 1.25×104 | 5.29×105 | 1.13×106 | 1.55×107 | 7.6×107 |
| Blank mixing | 3.31×103 | 8.15×104 | 2.53×106 | 1.76×107 | 4.35×108 | 9.38×109 |
| Fresh-keeping agent | 1.16×103 | 1.87×103 | 3.31×103 | 3.02×104 | 1.20×106 | 8.13×106 |
From the results, the preservative can effectively improve the sensory acceptability of the carp flesh compared with the red kojic rice wine and the blank mixed liquid.
In the preservative, the spice is added, so that the putrefaction speed of the carp and the fish can be effectively delayed, and the preservative has an obvious bacteriostatic and fresh-keeping effect. Thereby prolonging the shelf life of the carps and achieving the effect of fresh keeping.
In the preservative, due to the addition of the spice, obvious bacteriostatic and bactericidal substances exist, and the degradation of microorganisms on nutrient substances in food is effectively reduced. Thereby prolonging the shelf life of the carps and achieving the effect of fresh keeping.
The method is characterized in that the spice and the red yeast rice wine have strong positive synergistic effect on bacteriostasis by controlling the amount of the effective components, the red yeast rice wine can play a flavor effect while being synergistic with the spice, and the meat texture of fish meat is kept, and in addition, multiple tests show that the freshness preservation test of fish in different seasons has different freshness preservation periods, the freshness preservation period is prolonged to 15-20 days in low-temperature seasons due to the fact that the fish has less bacteria content in vivo and in vitro, and the freshness preservation period is 5-12 days in high-temperature seasons due to the fact that the fish has more bacteria content.
It should be noted that the above-mentioned embodiments are only for illustrating the technical solutions of the present invention and not for limiting, and although the present invention has been described in detail with reference to the preferred embodiments, it should be understood by those skilled in the art that modifications or equivalent substitutions may be made on the technical solutions of the present invention without departing from the spirit and scope of the technical solutions of the present invention, which should be covered by the claims of the present invention.
Claims (2)
1. A freshwater fish flesh preservative is characterized in that: measured by red yeast rice wine fermented at a constant temperature of 25 ℃ for 15 days, comprises,
mixing dark plums, nutmeg, clove and garlic in equal volume, and adopting an extracting solution extracted by red kojic rice wine, wherein the material-liquid ratio of the dark plum extracting solution is 67-72 g/L, the material-liquid ratio of the nutmeg extracting solution is 90-100 g/L, the material-liquid ratio of the clove extracting solution is 40-50 g/L, and the material-liquid ratio of the garlic extracting solution is 125-143 g/L;
the preparation method of the red yeast rice wine comprises the following steps,
soaking: soaking the sticky rice in clear water for 3.5-4.5 hours;
and (3) cooking: after being easily crushed by hand, the glutinous rice is put into a pot for cooking, and when the glutinous rice is cooked to be eighty percent cooked, the glutinous rice is taken out and cooled;
mixing: uniformly mixing red yeast rice and glutinous rice according to the ratio of 1: 5-1: 6, putting the mixture into an aseptic jar, and adding water, wherein the ratio of water to rice is 1.5: 1-2: 1;
fermentation: placing the jar in a thermostat at 25 deg.C, and fermenting for 15 days to obtain red rice wine;
the dark plum extract is extracted from dark plum, wherein the material-to-liquid ratio of total organic acid is 23-25 g/L; the extraction method of the dark plum extract comprises the steps of crushing and sieving dark plums, dissolving the crushed dark plums in the red yeast rice wine according to the liquid-material ratio of 14-15 ml/g, and extracting for 70-90 min by using an ultrasonic cleaner with the temperature of 55 ℃, the frequency of 40kHz and 200W;
the nutmeg extract is extracted from nutmeg, wherein the material-liquid ratio of nutmeg volatile oil is 26-29 g/L; the extraction method of the nutmeg extract comprises the steps of crushing and sieving nutmeg, dissolving the nutmeg in red yeast rice wine according to the liquid-material ratio of 10-11 ml/g, and extracting for 2-2.5 min by using an ultrasonic microwave instrument with the frequency of 40kHz and the power of 250-300W;
the clove extracting solution is extracted from clove, wherein the eugenol material-liquid ratio is 3.6-4.5 g/L; the extraction method of the clove extracting solution comprises the steps of crushing and sieving clove, dissolving the crushed clove in red kojic rice wine according to the liquid-material ratio of 20-25 ml/g, and extracting for 60-90 min by using an ultrasonic cleaner with the temperature of 55-65 ℃, the frequency of 40kHz and 200W;
the garlic extract is extracted from garlic, wherein the garlicin feed-liquid ratio is 0.36-0.42 g/L; the extraction method of the garlic extract comprises the steps of dissolving the garlic extract in red kojic rice wine according to the liquid-material ratio of 7-8 ml/g, and extracting for 20-30 min by using an ultrasonic cleaner with the temperature of 30-40 ℃, the frequency of 40kHz and the frequency of 48-72W.
2. A fresh-keeping method for freshwater fish flesh is characterized by comprising the following steps: which is to slaughter the freshwater fish, wash the killed freshwater fish with water, slice, rinse and drain the freshwater fish, soak the freshwater fish by the freshwater fish fresh-keeping agent of claim 1, package and refrigerate the freshwater fish, and comprises the following specific steps,
killing and cleaning fresh freshwater fish, and cutting the fish into fish slices with the thickness of 1-2 cm and smooth surfaces;
washing the fillets in precooled sterile water for 10-15 s, and draining at the temperature of 0-4 ℃;
soaking the drained fillets in a pre-cooled coating preservative for 30-60 min, wherein the volume ratio of the fillets to the freshwater fish preservative is 1: 3;
and (4) sealing, packaging and refrigerating the fillets soaked with the freshwater fish flesh preservative.
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