CN103053666B - Method for determining best ice temperature cold induction mode of fish flesh - Google Patents

Abstract

The invention provides a method for keeping the freshness of fish meat and increasing the taste of fish meat induced by ice temperature and cold, comprising the following steps: a) determination of the freezing point of fish meat; b) determination of the lethal temperature of fish; c) treatment of live fish before cold induction: Live fish are temporarily kept in water 3-5°C higher than the lethal temperature for more than 1 hour, and then slaughtered in an environment 3-5°C higher than the lethal temperature, and the internal organs are removed, and the fish is whole, segmented or sliced; d) Cold induction: After the temperature of the fish is lowered to its ice temperature zone, store it in the ice temperature zone for a period of time. The present invention also provides a method for determining the best ice-temperature-cold induction method for fish meat, which is to measure the freshness index K value and taste components on the basis of the above method, and evaluate the freshness and taste of fish meat with different cold induction methods, Determine the optimal cold induction modality. The method of the invention can obtain fresh and delicious fish meat, provides a reference for the selection of fish meat storage and processing methods, and has important guiding significance for improving fish sales quality, especially maintaining the freshness of processed aquatic products.

Description

A Method for Determining the Best Ice Temperature and Cold Induction Method for Fish Meat

technical field

The invention relates to the technical field of food preservation, in particular to a method for determining the best ice temperature and cold induction method for fish meat.

Background technique

Tilapias, also known as African crucian carp, belongs to the genus Tilapia of the family Cichlidae of the order Perciformes. Because of its omnivorous, adaptable, high reproductive rate, fast growth, Strong disease resistance has become the main cultured fish in the world, and has also become an important freshwater culture object in my country. Because tilapia has relatively high requirements on water temperature, and the living temperature range is 15°C to 35°C, the breeding scale is relatively large in areas with high temperatures such as Guangdong in my country. Tilapia is a freshwater farmed fish cultivated by key scientific research in the world's aquaculture industry, and is known as one of the main sources of animal protein in the future.

After being caught and killed, fish such as tilapia are still undergoing various complex changes in their bodies. From freshness immediately after death to spoilage, they generally go through three stages of stiffness, autolysis and spoilage. Usually, the freshness of fish before or during stiffness is good, and the freshness and quality of fish begin to decline during the autolysis stage. When protein is decomposed by enzymes, substances such as amino acids increase, which provides conditions for the reproduction of bacteria.

Utilize adenosine triphosphate (ATP) in fish muscle to decompose in the early postmortem period, through adenosine diphosphate (ADP), adenosine monophosphate (AMP), inosine monophosphate (IMP), inosine (HxR) , Hypoxanthine (Hx), etc., where the ratio of (HxR+Hx) to the sum of ATP and its degradation products is the freshness index K value:

Generally, K value ≤ 20% is used as an excellent freshness index (the quality standard for raw fish in Japan), and K value ≤ 60% is used as a freshness standard for processed raw materials.

It is generally believed that IMP is the main component of umami taste, and the production of IMP can enhance the umami taste of fish. When the concentration of IMP decreases, the flavor of fish gradually becomes unacceptable.

Free amino acids are the main components of non-volatile nitrogenous substances in taste substances. When some free amino acids exist in fish muscle at a high enough concentration, they can act on the flavor of fish independently of other components. For example, glycine and alanine produce sweetness, lysine produces sweet and bitter taste, histidine forms the "meaty aroma" of some seafood, and glutamic acid, aspartic acid and alanine have effects on umami. greater contribution.

Prolonging the storage time of tilapia and other aquatic products, maintaining the freshness of fish meat and increasing the taste of fish meat are of great significance to improve the quality of fish sales, especially to maintain the freshness of processed aquatic products.

In terms of fresh-keeping technology of aquatic products, the existing literature at home and abroad mainly focuses on ice storage and cold storage, and there are more and more reports on ice temperature technology in China. Ice temperature refers to the temperature range from 0°C to the freezing point of organisms, and ice temperature storage refers to storage in this temperature zone. The outstanding advantage of ice temperature storage compared with frozen storage is that it does not destroy cells; the outstanding advantage of ice temperature compared with refrigeration is that not only the activities of harmful microorganisms and the activities of various enzymes are inhibited, the storage period is extended, and the food quality can be improved. quality. Research on ice temperature storage of herring (Liang Qiong, Wan Jinqing, Wang Guoqiang. Herring slices[J]. Food Science, 2010(06):270-273.), flounder (Li Hui, Liu Lianfeng, Yang Bofeng, etc. Ice temperature The freshness and texture changes of flounder under fresh-keeping conditions [J].) and other studies have shown that ice-temperature storage conditions can more effectively inhibit the reproduction of microorganisms in aquatic products and prolong the storage period of food.

In recent years, Japanese scholars have particularly emphasized two aspects in ice temperature technology (Kunihiko Hattori. Ice temperature technology to improve food freshness and taste [C].), one is the importance of "biological defense response", and the other is to prevent Temperature variation technology, that is, constant temperature technology. The former means that the fresh food on the dining table can be regarded as alive from the perspective of cells. Take the dead fish with viscera processed for family consumption as an example. Slowly lower the temperature of the fish to the freezing temperature zone, and artificially convey to it the feeling that winter is approaching. As it approaches the freezing temperature, it seems that the fish itself predicts that it will be killed. Like freezing to death, the concentration of antifreeze (such as amino acids, etc.) in the cells increases, and after a certain period of time, the taste will become delicious. In addition, harmful microorganisms and pathogenic bacteria are significantly reduced due to the fact that the ice storage temperature range is in the low temperature range. This temperature range is also an area where enzymes, yeast, and lactic acid bacteria that improve the taste and flavor are very active. Therefore, no preservatives are added.

However, it is not to say that any fresh food or food material can increase the deliciousness of food due to the action of biological defense reactions and enzymes. Improving the effectiveness of the defense response of the organism by promoting it is one of the characteristics of ice temperature technology. For different food materials, in order to maximize the defense reaction of the organism, that is, the transformation of amino acids, the guidance of the cooling temperature is different, which is the biggest reason why the ice temperature effect is not easy to achieve: usually, for guiding the temperature to a higher temperature (About 10~5℃), even if rapid cooling is used, it seems that the ice temperature effect will not be greatly affected. However, from now on, the rate of temperature drop must be very slow, not impatient, and the so-called low-temperature domestication operation must be carried out to artificially pass on the feeling that winter is approaching to the creatures in nature, so that they can start to prepare for the arrival of winter physiologically. As a result, it can be said that the cold resistance has been added to the organism. However, for different food materials, it is not only necessary to cool down at a certain ratio, but also to set a gap jump time in the process seems to be an effective means. In addition, like some citrus, before entering the low-temperature field, there are some fruit and vegetable foods that may cause low-temperature damage in the temperature zone below +5°C. In this case, countermeasures to eliminate low-temperature damage should be found at the same time.

The Chinese journal "Food Science", Issue 06, 2010, published the paper "Study on Ice-temperature Storage of Herring Fillets", which put herring fillets at (-0.8±0.2), (-2.0±1.0) ℃ and (4.0 ±1.5) ℃ environment, take out regularly to measure its sensory, microbiological and physical and chemical indicators. At the end of the storage period, the total bacterial colonies of the samples under the three storage conditions were 1.17×10 ⁶ (day 11), 1.12×10 ⁶ CFU/g (day 8) and 1.2×10 ⁶ CFU/g (day 5); TVB-N values were 24.95 (day 20), 20.03mg/100g (day 15) and 20.39mg/100g (day 12); pH values were 6.96 (day 12), 6.59 (day 7) and 6.73 (day 8). The results showed that the herring fillets were close to spoilage on the 5th and 8th day of the experiment respectively under cold storage and slightly freezing conditions, while the herring fillets stored at ice temperature were close to spoilage on the 11th day.

The Chinese journal "Advances in Fishery Science", Issue 03, 2011, published a paper "Freshness and texture changes of flounder under ice temperature preservation conditions", researching the freshness index (pH) of flounder under ice temperature storage conditions value, K value, TVB-N, total number of bacteria) and texture characteristic parameters (sensory evaluation, tissue structure, breaking strength, etc.), and compared with the corresponding changes of refrigerated samples. The results showed that with the prolongation of storage time, the total number of colonies, K value and TVB-N increased, and the pH value decreased first and then increased, and the pH value of samples in the late storage period increased rapidly and was very high; the breaking strength decreased at the end of the shelf life. Small trend, the organizational structure is gradually deteriorating. Compared with refrigerated samples, ice-temperature storage conditions can more effectively inhibit the action of microorganisms in the flounder fish and prolong the storage period of flounder.

The ice temperature method of the above-mentioned aquatic products is to directly store fresh samples in the ice temperature range, but there is no report about the ice temperature after cold induction to maintain the freshness of fish meat and increase the taste of fish meat, and there is no effective determination The method of the optimal ice temperature and cold induction mode of fish meat has not yet been reported.

Contents of the invention

The purpose of the present invention is to aim at the deficiencies in the prior art, and provide a method for maintaining the freshness of fish meat and increasing the taste of fish meat induced by ice temperature.

The second object of the present invention is to provide a method for determining the optimal ice temperature and cold induction method for fish meat.

For realizing above-mentioned object, the technical scheme that the present invention takes is:

The invention relates to a method for maintaining the freshness of fish meat and increasing the taste of fish meat induced by ice temperature, which comprises the following steps:

a) Determination of the freezing point of fish meat;

b) determine the lethal temperature of fish;

c) Treatment of live fish before cold induction: temporarily raise the live fish in water 3-5°C higher than the lethal temperature for more than 1 hour, slaughter the fish in an environment 3-5°C higher than the lethal temperature, remove internal organs, and remove the whole fish , segment or slice;

d) Cold induction: After the temperature of the fish meat is lowered to its ice temperature zone, store it in the ice temperature zone for a period of time.

The specific method for determining the freezing point of fish in step a) is: slaughter the fish, remove the viscera, clean it, insert a temperature sensor such as a thermocouple or a thermal resistance into a place about 0.5 cm below the surface of the fish and fix it, and put it in -20 In the freezer room at ℃, the temperature collection interval is 10s to collect data once. After the experiment, the freezing curve is drawn and the freezing point of the fish is obtained.

The specific method for determining the lethal temperature of fish in step b) is: put live fish in an ecological tank for temporary breeding, cool down the water at a rate of 1°C/24h, manually observe and record the death time and corresponding water temperature every 2 hours , the death standard of the test fish is that after the operculum stops flapping and there is no response to the needle, the test fish is placed in natural water temperature and does not resume activities within 5 minutes. The experiment was repeated 3 times, and the highest temperature when dead fish appeared was selected as the lethal temperature of fish.

For realizing above-mentioned second purpose, the technical scheme that the present invention takes is:

A method for determining the optimal ice temperature and cold induction mode of fish, which comprises the following steps:

a) Determination of the freezing point of fish meat;

b) determine the lethal temperature of fish;

c) Treatment of live fish before cold induction: temporarily raise the live fish in water 3-5°C higher than the lethal temperature for more than 1 hour, slaughter the fish in an environment 3-5°C higher than the lethal temperature, remove internal organs, and remove the whole fish , segment or slice;

d) Cold induction: put the fish meat in a cold environment for cooling induction, the final temperature of the cold induction of the fish meat is its ice temperature zone, and store the fish meat in the ice temperature zone for a period of time;

e) Measure the freshness index K value and taste components, evaluate the freshness and taste of fish with different cold induction methods, and determine the best cold induction method.

The specific method for determining the freezing point of fish in step a) is: slaughter the fish, remove the viscera, clean it, insert a temperature sensor such as a thermocouple or a thermal resistance into a place about 0.5 cm below the surface of the fish and fix it, and put it into - In the freezing room at 20°C, the temperature collection interval is 10s to collect data once. After the experiment, the freezing curve is drawn and the freezing point of the fish is obtained.

The specific method for determining the lethal temperature of fish in step b) is: put live fish in an ecological tank for temporary breeding, cool down the water at a rate of 1°C/24h, manually observe and record the death time and corresponding water temperature every 2 hours , the death standard of the test fish is to put the test fish in the natural water temperature after the operculum stops fanning and there is no response to the acupuncture, and then the test fish does not resume activities within 5 minutes, repeat the experiment 3 times, and select the highest temperature when the dead fish appears , as the lethal temperature of fish.

The present invention has the advantage that:

1. The present invention provides a method for maintaining the freshness of fish meat and increasing the taste of fish meat induced by ice temperature and cooling, so that delicious fish meat can be obtained;

2. The method for keeping the freshness of fish meat and increasing the taste of fish meat induced by ice temperature and cold of the present invention is to temporarily raise live fish in water with a temperature 3-5°C higher than the lethal temperature for more than 1 hour, and in an environment with a temperature 3-5°C higher than the lethal temperature Slaughtering the medium-sized fish plays an important role in maintaining the taste components of the fish and ensuring the transformation of the taste components during the induction process of ice temperature;

3. It is known that for different food materials, in order to maximize the transformation of the biological defense response, that is, the amino acid, the guidance of the cooling temperature is different. This is the biggest reason why the ice temperature effect is not easy to achieve. The present invention is effective for tilapia. The initial cold induction temperature for cold induction is 15°C, which can achieve good cold induction effect;

4. The method of the present invention to determine the best way to induce ice-cold fish meat is to prolong the storage time of aquatic products such as tilapia, maintain its freshness and increase its taste, for improving the quality of fish sales, especially for maintaining the quality of processed aquatic products Freshness has important guiding significance.

Description of drawings

Accompanying drawing 1 is tilapia freezing curve.

Accompanying drawing 2 is the change of temperature of tilapia fillets under different cold induction conditions.

Accompanying drawing 3 is the standard product HPLC pattern of ATP and its degradation products.

Accompanying drawing 4 is the change of K value of tilapia fillets under different cold induction conditions.

Accompanying drawing 5 is the change of IMP and ATP of tilapia under different cold induction conditions.

Accompanying drawing 6 is the cooling curve of tilapia fillet 35h before.

Accompanying drawing 7 is the change of K value of tilapia fillet under 5 kinds of different initial temperature cold induction conditions.

Accompanying drawing 8 is the change of tilapia IMP and ATP under cold induction conditions of 5 different initial temperatures.

Detailed ways

The specific embodiments provided by the present invention will be described in detail below in conjunction with the accompanying drawings.

Example 1

1 Materials and methods

1.1 Materials

Tilapia: Purchased from the Guzong Road Vegetable Market in Lingang New City, Shanghai, weighing 500-600g/tail, oxygenated and quickly transported back to the laboratory after purchase.

1.2 Reagents and equipment

Reagents: Standards include Adenosine Triphosphate (ATP) and its compounds Adenosine Diphosphate (ADP), Adenosine Monophosphate (AMP), Inosine Monophosphate (IMP), Inosine ( Inosine, HxR), hypoxanthine (Hypoxanthine, Hx), ATP, ADP, IMP, Hx standard products are produced by Sigma Company, AMP standard products are produced by Japan TCI Company, HxR standard products are produced by German Dr. Ehrenstorfer Company. Methanol is HPLC grade (Sinopharm Group Chemical Reagent Co., Ltd.); Phosphate HPLC grade (Shanghai Anpu Scientific Instrument Company); experimental water is ultrapure water; perchloric acid (PCA), potassium hydroxide, sodium hydroxide, phosphoric acid , sulfosalicylic acid were analytically pure.

Equipment: Shimadzu LC-2010CHT high performance liquid chromatography; HITACHI L-8800 amino acid automatic analyzer; Agilent 34972A Agilent temperature acquisition instrument; Shanghai Yiheng Scientific Instrument Co., Ltd. constant temperature and humidity box BPS-100CL (-10~100 ℃), BPS-250CB (-40~100℃), LHS-100CA (-20~35℃); Changshu-Xueke IMS-50 automatic snowflake ice machine; Ningbo Xinzhi Biotechnology SB25-12DT ultrasonic machine; Fluko FA25 laboratory high-shear homogenizer; Shimadzu AUW320 electronic balance; Leici PHS-3C pH meter; Shanghai Yiheng Scientific Instrument DHG-9053A electric blast drying oven; Changsha Xiangyi H2050R refrigerated centrifuge; Teng GM-0.33A diaphragm vacuum pump and solvent filter; Qingdao Haier BCD-216SCM refrigerator.

The equipment used in the experiment was cleaned by an ultrasonic machine for 20 minutes, rinsed repeatedly with distilled water, then rinsed with ultrapure water for 2 to 3 times, and dried in a dryer for later use.

1.3 Test method

1.3.1 Determination of the freezing point of fish meat

Cut the fish into two pieces along the spine, insert a thermocouple about 0.5cm below the surface of the fish and fix it, put it into a freezer at -20°C, collect data once at a temperature collection interval of 10s, draw a freezing curve after the experiment and Find the tilapia freezing point.

1.3.2 Determining the lethal temperature of fish

Put 10 live fish in an ecological tank for temporary breeding, cool down the water at a rate of 1°C/24h, manually observe the time of death and the corresponding water temperature every 2 hours, and check the death standard of the experimental fish when the gill cover stops flapping And the acupuncture did not respond, and then the experimental fish was placed in natural water temperature, and did not resume activities within 5 minutes. The experiment was repeated 3 times, and the highest temperature when dead fish appeared was selected as the lethal temperature of fish.

1.3.3 Treatment of live fish before cold induction

Temporarily raise live fish in water 3-5°C higher than the lethal temperature for 1-2 hours, slaughter the fish in an environment 3-5°C higher than the lethal temperature, remove internal organs, slice into fish fillets, put them in fresh-keeping bags in a cold environment.

1.3.4 Cold induction - setting of cooling programs for different cold induction methods

Three different cold induction methods, temperature-controlled continuous cooling (mode Ⅰ), temperature-controlled step cooling (mode Ⅱ), and direct cooling with ice temperature (mode Ⅲ), respectively used 3 constant temperature and humidity chambers. The initial cooling temperature is 15°C to ensure that the tilapia remains fresh at this temperature.

Method Ⅰ, first stabilize the constant temperature and humidity box at 15°C for 2 hours, place the fish fillets in the stable temperature field in the center of the box, and the cooling program is as follows: 1°C drop per hour between 15°C→1°C, 1°C→ Drop 0.5°C per hour between -0.5°C; let the fish slowly adapt to the cold environment until it falls to the ice temperature zone, and maintain the ice temperature condition for a period of time.

Mode Ⅱ, first stabilize the constant temperature and humidity box at 15°C for 2 hours, then place the fish fillets in the steady state area of the temperature field in the center of the box. 15°C to start the cooling timer) and then directly drop to 4°C and keep the temperature unchanged, and after 10 hours (start the cooling timer from 10°C) and then directly drop to -0.5°C and maintain the ice temperature for a period of time.

Mode Ⅲ, first stabilize the constant temperature and humidity chamber at -0.5°C for 2 hours, put the fish fillets, and maintain the ice temperature condition for a period of time.

1.3.5 Detection of ATP and its related compounds

Refer to Yokoyama (Yokoyama Y, Sakaguchi M, et al. Change in concentration of ATP-related compounds in various tissues of oyster during ice storage[J]. Nippon Suisan Gakkaishi,1992,58(11):2125-2136.) and Qiu Weiqiang (Qiu Weiqiang, Chen Gang, Chen Shunsheng, etc. Simultaneous detection of 6 ATP-related compounds in aquatic products by ion-pair reversed-phase high-performance liquid chromatography [J]. Fisheries Journal, 2011, (11): 1745-1752.), abbreviated There are changes.

(1) Extraction of ATP and related compounds (ADP, AMP, IMP, HxR, Hx) of fish fillets

Take fresh fish fillets and samples after cold treatment, chop them up quickly, put 5g into a centrifuge tube, add 10ml of pre-cooled 10% perchloric acid (PCA) solution and beat for 2min. After homogenization, freeze and centrifuge at 10000r/min for 15min, and take the supernatant. The precipitate was washed with pre-cooled 5% PCA, the supernatant was obtained by centrifugation, and the operation was repeated once. Combine the supernatant, first use 10mol/L KOH solution and then use 1mol/L KOH solution to adjust the pH value to 6.5, after standing for 30min, transfer the supernatant to a 50mL volumetric flask, dilute to volume with ultrapure water, shake well, After filtering with a 0.45 μm microporous membrane, wait for the determination. The whole process was operated at 0-4°C.

(2) High performance liquid chromatography (HPLC) detection chromatographic conditions

Chromatography (HPLC) conditions: GL Sciences Inertsil ODS-SP C18 (4.6×250mm, 5μm) liquid chromatography column; guard column core Inertsil ODS-SP (4×10mm, 5μm); mobile phase: A is 0.05mol/ L Potassium dihydrogen phosphate and dipotassium hydrogen phosphate (1:1) solution, adjusted to pH 6.5 with phosphoric acid, B is methanol solution; isocratic elution; flow rate: 1mL/min; column temperature: 28°C; injection volume : 10μL; detection wavelength: 254nm.

1.3.6 Detection of free amino acids

Refer to Deng Jiechun (Deng Jiechun, Wang Xichang, Liu Yuan. Study on the difference of taste components between puffer puffer obscurus and puffer red fin [J]. Food Industry Science and Technology, 2010, (3): 106-108.) and Xue Song (Xue Song, Wan Jin Qing. The effect of ice temperature storage on the freshness and free amino acid of chicken meat [J]. Jiangsu Agricultural Science, 2010(6): 411-413.) The method for the determination of free amino acid has been slightly modified.

(1) Extraction of free amino acids

Weigh 2 g of fish respectively, add 10 mL of 5% (w/v) sulfosalicylic acid, precipitate for 2 hours to precipitate the protein, absorb 6 mL of supernatant and centrifuge with 10000 r/min refrigerated centrifuge for 15 min, take 3 mL of supernatant, and use A certain amount of NaOH solution was used to adjust the pH to about 2.0, and the volume was adjusted to 12mL, filtered through a 0.45μm micropore, and put into a sample tray for measurement.

(2) Analysis conditions

L-8800 amino acid automatic analyzer, the sample analysis period is 53min.

Chromatographic column: 4.6×150mm, 7μm;

Column temperature: 50°C;

Flow rate 1: 0.4ml/min, flow rate 2: 0.35ml/min;

Mobile phase: buffer of citric acid, sodium citrate and ninhydrin.

2 Analysis of experimental results

2.1 Determination of freezing point

The freezing curve of tilapia is shown in Figure 1. The freezing point of tilapia is about -1.2°C. Therefore, it can be considered that when the storage temperature of tilapia fillets drops to freezing point and then continues to drop, the cells of the fillets begin to crystallize and destroy the cell structure. According to the measured freezing point of tilapia fillets, the experiment in this paper is to keep the temperature of the tilapia fillets in the range of (-0.5±0.3) °C after dropping the tilapia fillets to the ice temperature zone.

2.2 Determination of fish lethal temperature

After testing, it was determined that the lethal temperature of tilapia was 12°C.

2.3 Changes in temperature of fish fillets

The cooling curve of tilapia fillets for the first 30 hours is shown in Figure 2. The results show that the cooling curve of mode Ⅰ shows a uniform and continuous downward trend, and the cooling curve of mode Ⅱ shows a gradient decline. The two methods basically reach the ice temperate zone at about 17 hours. Mode Ⅲ is to put it directly into the ice temperature environment, and the falling speed is faster, and it will fall to the ice temperature zone in about 3 hours.

2.4 Changes in K value

Figure 3 is a high-performance liquid chromatogram containing 6 kinds of standard products ATP, ADP, AMP, IMP, HxR, Hx, ATP and its degradation products can be effectively separated within 18 minutes under this chromatographic condition, and the reproducibility is good . The freshness index K value is a biochemical index that reflects the initial freshness change and flavor of aquatic products (Wu Chengye, Ye Mei, Wang Qin, etc. Study on freshness changes of several freshwater fish during frozen storage). The smaller the K value, the better the freshness, and the larger the K value, the worse the freshness. Many scholars have studied the relationship between K value and freshness through aquatic products such as sardines, horse mackerel, conger eel, and large yellow croaker. K value is a generally accepted indicator for evaluating the early freshness of fish. Below 10%. The K value of raw fish fillets is less than 20% of the first-grade freshness, 20%-40% is the second-grade freshness, and 60%-80% is the initial spoilage fish. ADP, AMP, IMP, HxR, and Hx are produced during the degradation of ATP. The ratio of HxR+Hx to the sum of ATP and its degradation products is the K value, which can be expressed by the following formula:

By calculation, the changes in the K value of tilapia under the three cold induction conditions are shown in Figure 4. It can be seen from Figure 4 that within 10 hours, the K values of the three cold induction methods have little difference. From 10 to 53 hours, the K values of the three methods all showed an upward trend, and the method III was significantly slower than the other two. After 53h, the effects of different cold induction methods on the K value showed significant differences. At 53h, the K value of method Ⅰ was 16.2%, which could still be eaten as raw fish. At 65h, the K value rose to 25.6%, exceeding a 20% of the freshness of grade; in mode II, the K value was 18.3% at 65h, and the K value just reached 20% at 90h. In mode III, the K value was 11.7% at 65 hours, and only 13.8% at 90 hours, much lower than the former two induction methods. Until 126 hours, the K value was 22.2%. Different cold induction methods have a greater impact on the K value. The possible reason is that the average temperature of the fish in the same period of time is that the mode I is greater than the mode II, and the mode II is greater than the mode III, which affects the enzymes of ATP and its related compounds. activity, leading to different changes in K values. During the whole experiment, the freshness level of the three kinds of cooling methods was kept relatively high, and the K value was lower than 40% at 138h.

2.5 Nucleotide IMP and ATP analysis

Inosinic acid (IMP), degraded from adenosine triphosphate (ATP), is the main taste substance of nucleotides. As a new type of food additive, it has been widely used in food seasoning. The mixture of IMP and glutamic acid sodium salt has a very significant synergistic effect. Experiments show that if the two are mixed in a certain proportion, the umami taste will be greater than their own umami taste (Li Huifang, Chen Guohong, Wu Xinsheng, etc. Animal muscle inosinic acid research Progress [J]. Zoology and Animal Medicine, 1999,16(4):6-7.). IMP is a flavor enhancer with strong umami taste (Qi Xiaoyu, Li Yan, Zhou Peigen. Changes of ATP degradation products and freshness evaluation of Macrobrachium japonica during ice storage). With the decomposition of ATP, IMP shows an upward trend. It can be said that the highest concentration is reached within 1~2 days and maintained for a period of time, which is consistent with the experimental results. It can be seen from Figure 5 that the ATP content of the three cold induction methods showed a rapid decline trend in the first 20 hours, and the decline rate reached 92%, 93%, and 93%, respectively. The rapid degradation of ATP is related to highly active ATPase, Wataba et al. (Watabe S, Ushio H, Iwamoto M, et al. Temperature-dependency of rigor-mort is of fish muscle: myofibrillar Mg ²⁺ -ATPase activity and Ca ²⁺ uptake by sarcoplasmic reticulum [J]. J Food Sci, 1989,54:1107-1115.) believes that at low temperature, the calcium absorption capacity of the sarcoplasmic reticulum decreases, the calcium concentration in the myofibrils increases, and calcium ions activate myogenic Fiber Mg ²⁺ -ATPase, accelerates the degradation of ATP. The IMP content showed an upward trend and reached the peak before 20h, and then showed a downward trend. At 17h, the maximum IMP content measured in modes I, II, and III were 166.43mg/100g, 165.62mg/100g, and 167.75mg/100g, which were 30%, 28%, and 5% higher than those at 10h, respectively. 27%, 32%, and 7.8% more than those at 39h, and the IMP of the three methods were all at a higher level at 17-27h. After 50 hours of IMP content, there was a significant difference. The IMP decreased significantly in mode Ⅰ, followed by mode Ⅱ, and the slowest decline in mode Ⅲ. The experimental results showed that in the late stage of cold induction, different cooling methods affected the decomposition rate of IMP. The degradation of IMP in mode Ⅰ was significantly faster than that in mode Ⅱ in the late stage of cold induction, and the degradation of IMP in mode Ⅲ was relatively slow, and the IMP content was still higher than that just after slaughtering at 138 hours.

2.6 Analysis of main free amino acids in different cold induction methods

Table 1 Changes of free amino acids in the taste of tilapia fillets under different induction methods

Note: Ⅰ—continuous cooling method with temperature control, Ⅱ—step cooling method with temperature control, Ⅲ—direct cooling method with ice temperature, TFAA—total amount of free amino acids (including 17 kinds of amino acids aspartic acid, threonine, serine, glutamine Acid, glycine, alanine, cystine, valine, methionine, isoleucine, leucine, tyrosine, phenylalanine, lysine, histidine, arginine, proline acid).

Table 1 lists the changes of the free amino acid content and total free amino acid content of eight flavors of tilapia fillets in the first 65 hours. Since the IMP of the three cold-induced methods decreased significantly after 65h, the subsequent changes were not listed in the table.

Compared with 0h after slaughter, the TFAA of ways Ⅰ, Ⅱ, and Ⅲ increased by 35.96%, 21.86%, and 20.03% at 17h; Increased by 49.61%, 29.7%, and 25.2%. Generally speaking, the TFAA content of the three methods showed an upward trend, and the TFAA content of method I increased significantly, which was higher than that of methods II and III. The delicious taste in fish meat is mainly determined by amino acids such as aspartic acid, glycine, glutamic acid, alanine and arginine, and the results show that aspartic acid, glycine, glutamic acid, alanine The four kinds of umami-tasting free amino acids account for 58.04% of TFAA, and the influence of these ingredients on the flavor of fish meat is obvious.

The content of glycine and alanine in the main taste free amino acids of tilapia is relatively high. Glycine is the main taste component of seafood products, and it has a synergistic effect with other umami substances such as glutamic acid and inosinic acid. . Methods Ⅰ, Ⅱ, Ⅲ, three kinds of cold induction methods, compared with 0h after fish slaughter, at 17h, the content of glycine increased by 47.07%, 13.81%, 7.38%; at 27h, increased by 51.58%, 26.25% , 19.54%; increased by 76.16%, 27.24%, 26.44% at 65h. The increase in mode Ⅰ was more obvious, exceeding the threshold of 130mg/100g at 17h, and the increase in mode Ⅱ and Ⅲ was relatively slow after 27h. Alanine is a sweet amino acid with a slightly bitter taste, and it can also bring out the umami components of fish and shellfish. In mode I, the increase of alanine reached 14.04% in 27 hours, in mode II it only increased by 4.8%, and in mode III there was basically no increase and change. The content of alanine fluctuated around 20mg/100g throughout the process, and the trend of change was relatively flat. The threshold value of glutamic acid in fish meat is mostly below 0.03%, but it has a synergistic effect with the IMP accumulated in fish meat, and even if the content is below the threshold value, it can still produce umami taste. Ⅰ, Ⅱ, Ⅲ, the content of glutamic acid in the three methods showed an increasing trend, which increased by 22%, 26% and 3% respectively at 27h compared with 0h. The threshold value of amino acid is 5mg/100g. Histidine is the characteristic that causes the "meaty aroma" of some seafood (Shen Yuexin. Aquatic Food Science [M]. China Agricultural Press. 2001:31-35.), the content of histidine at 27h is 17.17mg /100g, 16.70mg/100g, 15.96mg/100g, basically close to the threshold value of 20mg/100g, which increased by 86.26%, 81.21%, and 73.14% respectively compared with 0h.

Arginine is a bitter amino acid with a small content. It does not have a bitter taste in aquatic products, but instead increases the freshness effect. Its effect on the persistence, complexity and richness of the taste cannot be ignored (Li Huifang, Chen Guohong, Wu Xinsheng, etc. . Research progress of animal muscle inosinic acid [J]. Zoology and Veterinary Medicine, 1999,16(4):6-7.). The bitter amino acid methionine fluctuates around 10mg/100g, the change trend is not obvious and is lower than the threshold of 30mg/100g. It is an indispensable taste component for the unique flavor of sea urchin. Studies have found that a small amount of methionine can improve the taste of MSG (Deng Jiechun, Wang Xichang, Liu Yuan. Study on the Differences in Taste Components between Fugu obscurus and Fugu obscurus [J]. Food Industry Science and Technology, 2010, (3): 106-108. ). Isoleucine is also a bitter amino acid, the content of which is much lower than their threshold value of 90mg/100g in the whole cooling process, and there are signs of reduction; aspartic acid with umami taste is also slightly reduced, but because of Its content is low and has limited impact on the overall taste.

3 Conclusion

By using three different cold induction methods to conduct cold induction experiments on tilapia fillets, the following conclusions were drawn:

(1) The K values of cold induction methods Ⅰ, Ⅱ, and Ⅲ were within 60h, 90h, and 120h respectively, and none of them exceeded 20%, which met the eating standards of fresh fish fillets; different cold induction methods had different influences on K values. Hysteresis, after 53 hours, the K value of method Ⅰ increases rapidly, which means that the freshness decreases rapidly.

(2) The nucleotide (IMP) of cold induction method III grows fast and degrades slowly in the later stage. After slaughtering tilapia fillets, they can quickly drop to the ice temperature zone, which can significantly delay the decline of IMP, which is conducive to the retention of its corresponding taste .

(3) In the same period of time, the cold induction method Ⅰ can significantly increase the total amount of free amino acids in tilapia fillets, especially the content of glycine, which exceeds the threshold value of 130mg/100g at 17h.

(4) Considering the changes of K value, nucleotide (IMP) and main taste free amino acids under different conditions, the tilapia fillets induced by method I were the most delicious during 17-27h.

Example 2

1 Materials and methods

1.1 Materials and equipment

Tilapia: In August 2012, tilapia was purchased from the Guzong Road Vegetable Market in Lingang New Town, Shanghai, with a weight of 500-600g/tail. After purchase, it was oxygenated and quickly transported back to the laboratory. Store the live fish in a preservation box for 1~2 hours to prevent them from struggling, then beat the live fish to death, slice them into fish fillets, put them in fresh-keeping bags and place them in a cold environment; adenosine triphosphate (ATP), diphosphate Adenosine (ADP), inosinic acid (IMP), hypoxanthine (Hx): Sigma Company; adenosine monophosphate (AMP): Japan TCI Company; inosine (HxR): German Dr.Ehrenstorfer Company; dihydrogen phosphate Potassium, potassium dihydrogen phosphate: Shanghai Anpu Scientific Instrument Co., chromatographically pure; ultrapure water, perchloric acid (PCA), potassium hydroxide, sodium hydroxide, phosphoric acid, sulfosalicylic acid: Sinopharm Chemical Reagent Co., Ltd. , analytically pure; Methanol: Sinopharm Chemical Reagent Co., Ltd., chromatographically pure.

LC-2010CHT high-performance liquid chromatography, AUW320 electronic balance: Shimadzu Corporation; L-8800 amino acid automatic analyzer: HITACHI Corporation; Agilent-34972A temperature acquisition instrument: Agilent Corporation; BPS-250CB (-20~100℃) Constant temperature and humidity box, DHG-9053A electric blast drying oven: Shanghai Yiheng Scientific Instrument Co., Ltd.; IMS-50 automatic snowflake ice machine: Changshu Snow Science Co., Ltd.; SB25-12DT ultrasonic machine: Ningbo Xinzhi Biotechnology; FA25 Homogenizer: Fluko Company; PHS-3C pH Meter: Shanghai Precision Scientific Instrument Co., Ltd.; H2050R Refrigerated Centrifuge: Changsha Xiangyi Co., Ltd.; GM-0.33A Diaphragm Vacuum Pump and Solvent Filter: Tianjin Jinteng Experimental Equipment Co., Ltd.; Haier BCD-216SCM refrigerator: Qingdao Haier Company. The equipment used in the experiment was cleaned by an ultrasonic machine for 20 minutes, rinsed repeatedly with distilled water, then rinsed with ultrapure water for 2 to 3 times, and dried in a dryer for later use.

1.2 Experimental method

1.2.1 Determination of freezing point

Cut the fish into two pieces along the spine, insert a thermocouple about 0.5cm below the surface of the fish and fix it, put it into a freezer at -20°C, collect data once at a temperature collection interval of 10s, draw a freezing curve after the experiment and Find the tilapia freezing point.

1.2.2 Setting of the cooling program of the cold induction method with different initial cold induction temperatures

Five different initial temperatures, respectively 25, 20, 15, 10, and 5°C, all adopt temperature control to cool down. The cooling program is shown in Table 2.

Table 2 Setting of cooling program

Cool down at the same cooling rate, let the fish slowly adapt to the cold environment until it falls to the ice temperature zone, and maintain the ice temperature condition for a period of time.

1.2.3 Detection of ATP and its related compounds

Refer to Yokoyama (Yokoyama Y, Sakaguchi M, Kawai F, et al. Change in concentration of ATP-related compounds in various issues of oyster during ice storage[J]. Nippon Suisan Gakkaishi,1992,58(11):2125-2136.) method, slightly modified. Extraction of fish fillet ATP and related compounds: Take fresh fish fillets and samples after cold treatment, quickly chop them, take 5g and put them into a centrifuge tube, add 10mL of pre-cooled 10% perchloric acid (PCA) solution to beat for 2min. After homogenization, freeze and centrifuge at 10000r/min for 15min, and take the supernatant. The precipitate was washed with pre-cooled 5% PCA, the supernatant was obtained by centrifugation, and the operation was repeated once. Combine the supernatants, first use 10mol/L KOH solution to adjust the pH value, when it is close to the required pH value, use 1mol/L KOH solution to fine-tune the pH value to 6.5, and after standing for 30min, transfer the supernatant to 50mL volumetric flask, and dilute to volume with ultrapure water, shake well, filter with 0.45μm microporous membrane and wait for determination. The whole process was operated at 0-4°C.

High-performance liquid chromatography (HPLC) detection chromatographic conditions: GL Sciences Inertsil ODS-SP C ₁₈ (4.6mm×250mm, 5μm) liquid chromatography column; guard column core Inertsil ODS-SP (4mm×10mm, 5μm); flow Phase: A is 0.05mol/L potassium dihydrogen phosphate and dipotassium hydrogen phosphate (1:1) solution, adjusted to pH 6.5 with phosphoric acid, B is methanol solution; isocratic elution; flow rate: 1mL/min; column temperature : 28°C; injection volume: 10 μL; detection wavelength: 254nm.

1.2.4 Detection of free amino acids

Refer to the free amino acid determination method of Deng Jiechun (Deng Jiechun, Wang Xichang, Liu Yuan. Study on the difference of taste components between puffer puffer obscura and puffer red fin [J]. Food Industry Science and Technology, 2010,31(3):106-108.), abbreviated There are changes. Extraction of free amino acids: Weigh 2g of fish meat respectively, add 10mL of 5% (w/v) sulfosalicylic acid, precipitate for 2h to precipitate the protein, absorb 6mL supernatant and centrifuge for 15min in a refrigerated centrifuge at 10000r/min, take 3mL For the supernatant, use a certain amount of NaOH solution to adjust the pH to about 2.0, set the volume to 12mL, filter through 0.45μm micropores, put it into the sample tray and use it for measurement.

Analysis conditions: L-8800 amino acid automatic analyzer, sample analysis cycle 53min. Chromatographic column (4.6mm×150mm, 7μm); column temperature: 50°C; channel 1 flow rate: 0.4mL/min, channel 2 flow rate: 0.35mL/min. Mobile phase: citric acid and sodium citrate mixed buffer with pH values of 3.3, 3.2, 4.0, and 4.9 and a buffer with a concentration of 4% ninhydrin.

2 Results and discussion

2.1 Determination of freezing point

The freezing curve of tilapia is shown in Figure 1. The freezing point of tilapia is about -1.2°C. Therefore, it can be considered that when the storage temperature of tilapia fillets drops to freezing point and then continues to drop, the cells of the fillets begin to crystallize and destroy the cell structure. According to the measured freezing point of tilapia fillets, the experiment in this paper is to keep the temperature of the tilapia fillets in the range of (-0.5±0.3) °C after dropping the tilapia fillets to the ice temperature zone.

2.2 Changes in temperature of fish fillets

The cooling curve of tilapia fillets for the first 35 hours is shown in Figure 6. The results showed that the cooling curves of modes Ⅰ, Ⅱ, Ⅲ, Ⅳ and Ⅴ showed a continuous cooling trend, and reached the ice temperate zone at about 27, 22, 17, 12, and 7 hours respectively, and maintained in the ice temperate zone for a period of time. The temperature of the fish fillet is (-0.5±0.3) ℃.

2.3 Changes in K value of freshness index

Through calculation, the change of K value of tilapia is shown in Figure 7 under the five cold induction methods with different initial temperatures of Ⅰ, Ⅱ, Ⅲ, Ⅳ and Ⅴ. It can be seen from Fig. 7 that the influence of five cold induction methods with different initial temperatures on the K value of tilapia fillets, the K value in the process of cooling, with the longer storage time, all showed an upward trend, and the K value of fresh fish fillets The value is 2.27%. The K value of mode Ⅰ and Ⅱ increased rapidly, followed by mode Ⅲ, and the growth rate of mode Ⅳ and Ⅴ was slow, showing a gentle upward trend. After 27 hours of method I induction, the K value just reached 20%, which was still in the first-grade freshness range; after 28-70 hours, the K value exceeded 40%, and the fish fillets were in the second-grade freshness stage; at 171 hours, the K value was 46.1%, not yet corrupted. In mode Ⅱ, at 30 hours, the K value exceeds 20% of the first-class freshness index, at 70-142 hours, the K value exceeds 30%, and at 142 hours, it is close to the second-class freshness index. Method Ⅲ was still within the range of the first grade freshness index at 50h, until 161h, the K value was 36.7%, within the range of the second grade freshness index. Mode Ⅳ and mode Ⅴ reached the first-level freshness index at 108h and 127h respectively, and did not exceed 30% at 150h, maintaining a relatively high freshness level.

2.4 Nucleotide IMP and ATP analysis

Inosinic acid (IMP), degraded from adenosine triphosphate (ATP), is the main taste substance of nucleotides and has been widely used in food seasoning. IMP is a flavor enhancer with strong umami taste. With the decomposition of ATP, IMP tends to increase. Generally speaking, it reaches the highest concentration within 1~2 days and maintains it for a period of time, which is consistent with the experimental results. The rapid degradation of ATP is related to the high activity of ATPase, Wataba et al. (Watabe S, Ushio H, Iwamoto M, et al. [J]. J Food Sci, 1989,54:1107-1115.) believes that at low temperature, the calcium absorption capacity of the sarcoplasmic network structure decreases, the calcium concentration in the myofibril increases, and calcium ions activate the myofibril Mg ²⁺ -ATPase, which accelerates the degradation of ATP. The changes of tilapia IMP and ATP under cold induction conditions at different initial temperatures are shown in Figure 8. It can be seen from Figure 8 that within 40 hours, IMP all peaked and degraded at different speeds after 40 hours. The degradation of mode Ⅰ and mode Ⅱ The speed is faster, and it is lower than the IMP value at 50h when it was just slaughtered. Modes Ⅲ and Ⅳ were lower than the IMP value of 2.62 μmol/g when just slaughtered, and the time was 89 and 150 hours respectively. The IMP content of pattern Ⅴ has always been at a relatively high level, and it was still higher than the level just slaughtered after the end of the experiment.

2.5 Analysis of free amino acids

Table 3 Composition of free amino acids in the taste of tilapia fillets (mg/100g)

Note: Ⅰ: 20°C is the cold induction method of the initial temperature, Ⅱ: 15°C is the cold induction method of the initial temperature, Ⅲ: 10°C is the cold induction method of the initial temperature, Ⅳ: 15°C is the cold induction method of the initial temperature, Ⅴ: 5°C is the cold induction method with the initial temperature; TFAA includes 17 kinds of amino acids (aspartic acid, threonine, serine, glutamic acid, glycine, alanine, cystine, valine, methionine, iso Leucine, Leucine, Tyrosine, Phenylalanine, Lysine, Histidine, Arginine, Proline).

Table 3 lists the changes of six flavors of free amino acids and the total amount of free amino acids in tilapia. As far as TFAA is concerned, both modes Ⅰ and Ⅱ have a flat growth trend, modes Ⅲ and Ⅳ have relatively obvious increases, and mode Ⅴ has the slowest growth rate. Compared with the 0h after slaughter, at 27h, they increased by 10.81%, 10.31%, 21.46%, 14.87%, 4.97%, respectively; %; at 123h, they increased by 16.43%, 19.19%, 32.63%, 25.36%, and 14.91%, respectively. The TFAA content of the above five cold-induced modes with different initial temperatures all showed an upward trend, and the mode Ⅲ and Ⅳ increased significantly, and the mode Ⅲ was greater than the mode Ⅴ, and all were higher than the modes Ⅰ, Ⅱ and Ⅴ. The delicious taste of fish meat is mainly determined by amino acids such as glycine, glutamic acid, alanine, leucine, isoleucine, and histidine. The experimental results show that in freshly slaughtered tilapia fillets, asparagine The 4 main flavor free amino acids, acid, glutamic acid, glycine and alanine, accounted for 57.28% of the measured TFAA, and the influence of these components on the taste of fish is very obvious.

The content of free amino acids in fish is lower than that of shellfish, but some free amino acids can exist in fish meat in sufficient and high concentrations to have an effect on the flavor of fish. Among the taste free amino acids in tilapia fillets, the contents of glycine and alanine are relatively high, among which glycine is the main taste substance of seafood products, and it has a synergistic effect with umami substances such as inosinic acid and glutamic acid . Compared with 5 cold induction methods with different initial temperatures at 27h, the content of glycine in mode Ⅲ was the highest, which was close to the threshold value of glycine 130mg/100g, which was 34.18% higher than that of the original, and the mode Ⅰ, Ⅱ, and Ⅳ increased by 20.98% and 18.65% respectively. %, 28.10%, and mode V has the smallest growth potential. Fish and shellfish contain glutamic acid. The threshold value of glutamic acid in fish meat is below 0.03%, but it has a multiplied effect with inosinic acid in postmortem muscle, so even if the content is above the threshold value, it can still produce umami taste ^{[17 ]} . The five cooling methods with glutamic acid basically exceeded the threshold value of 5mg/100g at 27h, and the increases were 16.04%, 20.37%, 23.12%, 23.74%, and 2.89% compared with the original. The increases at 75 hours were 5.74%, 30.30%, 37.07%, 18.65%, and 10.53%, respectively, and the increase of glutamic acid in mode Ⅲ was relatively obvious. Alanine is a sweet amino acid that contributes to the umami of fish. The content of alanine in the five methods fluctuated around 21mg/100g, and the change was relatively gentle. Histidine is the characteristic that causes the meaty aroma of certain seafood, and the increase in mode Ⅰ, Ⅱ, Ⅲ is more significant, and the increase is 69.41%, 44.50%, 95.22% at 27h compared with 0h, and the mode Ⅳ, Ⅴ basically does not increase.

Leucine and isoleucine are bitter amino acids. The results of the study found that the content of leucine and methionine was far below their thresholds of 190mg/100g and 90mg/100g, and there was a tendency to decrease during the cooling process. In addition, the content of arginine and methionine are also bitter amino acids, and arginine plays an important role in the persistence and complexity of the taste, and methionine is an indispensable flavor component of sea urchin.

3 Conclusion

3.1 Cold induction methods Ⅰ, Ⅱ, Ⅲ, Ⅳ, Ⅴ have different effects on the freshness of tilapia. The lower the initial induction temperature, the slower the increase of K value and the better the freshness, otherwise the faster the increase of K value, the lower the freshness.

3.2 For cold induction methods Ⅰ, Ⅱ, Ⅲ, Ⅳ, and Ⅴ, the IMP content peaked within 40 hours, and at the same time ATP degraded rapidly.

3.3 The total amount of free amino acids increased in cold induction methods Ⅰ, Ⅱ, Ⅲ, Ⅳ, and Ⅴ, and the degree of increase was different for different induction methods. The study found that the initial temperature of cold induction was 15°C, and the amount of free amino acids increased significantly.

The above is only a preferred embodiment of the present invention, it should be pointed out that for those of ordinary skill in the art, without departing from the method of the present invention, some improvements and supplements can also be made, and these improvements and supplements should also be considered Be the protection scope of the present invention.

Claims (6)

1. a kind of ice temperature cold induction keeps the method for freshness of fish meat and increases the taste of meat of fish, it is characterized in that, it comprises the following steps: a) Determination of the freezing point of fish meat; b) determine the lethal temperature of fish; c) Treatment of live fish before cold induction: temporarily raise the live fish in water 3-5°C higher than the lethal temperature for more than 1 hour, slaughter the fish in an environment 3-5°C higher than the lethal temperature, remove internal organs, and remove the whole fish , segment or slice; d) Cold induction: After the temperature of the fish meat is lowered to its ice temperature zone, it is stored in the ice temperature zone for a period of time; wherein, the above-mentioned lowering the temperature of the fish meat to its ice temperature zone refers to placing the fish meat in a cold environment for temperature control and continuous cooling induction or Temperature-controlled gradient cooling induction, the initial temperature of fish meat cold induction is 3-5°C higher than the lethal temperature, and the end temperature of fish meat cold induction is in its ice temperature zone. The procedure for temperature-controlled continuous cooling induction is: at the initial temperature→ Decrease 1°C per hour between 1°C and 0.5°C per hour between 1°C→freezing point of fish meat. The temperature control step-down temperature induction procedure is as follows: drop directly from the initial temperature to 10°C and keep the temperature constant. The initial temperature starts to cool down for 5 hours, then directly drops to 4°C and keeps the temperature constant, and starts to cool down from 10°C for 10 hours, and then directly drops to the freezing point of the fish. 2. The method according to claim 1, characterized in that the method for determining the freezing point of fish in step a) is as follows: slaughter the fish, remove the viscera, clean it, and insert temperature sensors such as thermocouples or thermal resistors The fish body was fixed at about 0.5cm below the surface, and placed in a freezer at -20°C. The temperature collection interval was 10s to collect data once. After the experiment, the freezing curve was drawn and the freezing point of the fish was obtained. 3. The method according to claim 1, characterized in that the specific method for determining the lethal temperature of fish in step b) is: put the live fish in an ecological tank for temporary breeding, and set the temperature at a rate of 1°C/24h. Cool down the water, manually observe the time of death and the corresponding water temperature every 2 hours, and check the death standard of the experimental fish. After the gill cover stops flapping and there is no response to the needle, the experimental fish is placed in the natural water temperature and does not recover within 5 minutes. Activity, repeat the experiment 3 times, select the highest temperature when dead fish appear, as the lethal temperature of fish. 4. A method for determining the optimal ice temperature and cold induction mode of fish, is characterized in that it comprises the following steps: a) Determination of the freezing point of fish meat; b) determine the lethal temperature of fish; c) Treatment of live fish before cold induction: temporarily raise live fish in water 3-5°C higher than the lethal temperature for more than 1 hour, slaughter the fish in an environment 3-5°C higher than the lethal temperature, remove internal organs, , segment or slice; d) Cold induction: After the temperature of the fish meat is lowered to its ice temperature zone, it is stored in the ice temperature zone for a period of time; wherein, the above-mentioned lowering the temperature of the fish meat to its ice temperature zone refers to placing the fish meat in a cold environment for temperature control and continuous cooling induction or Temperature-controlled gradient cooling induction, the initial temperature of fish meat cold induction is 3-5°C higher than the lethal temperature, and the end temperature of fish meat cold induction is in its ice temperature zone. The procedure for temperature-controlled continuous cooling induction is: at the initial temperature→ Decrease 1°C per hour between 1°C and 0.5°C per hour between 1°C→freezing point of fish meat. The temperature control step-down temperature induction procedure is as follows: drop directly from the initial temperature to 10°C and keep the temperature constant. The initial temperature starts to cool down for 5 hours, then directly drops to 4°C and keeps the temperature unchanged, starts to cool down from 10°C for 10 hours, and then directly drops to the freezing point of the fish; e) Measure the freshness index K value and taste components, evaluate the freshness and taste of fish with different cold induction methods, and determine the best cold induction method. 5. The method according to claim 4, characterized in that, the specific method for determining the freezing point of fish meat in step a) is: slaughter the fish, remove the viscera, clean it, and place temperature sensors such as thermocouples or thermal resistors, Insert it at about 0.5cm below the surface of the fish body and fix it, put it into a freezer at -20°C, collect data at a temperature collection interval of 10s, draw a freezing curve after the experiment and get the freezing point of the fish. 6. The method according to claim 4, characterized in that the specific method for determining the lethal temperature of fish in step b) is: put the live fish in an ecological tank for temporary breeding, and set the temperature at a rate of 1°C/24h. Cool down the water, manually observe the time of death and the corresponding water temperature every 2 hours, and check the death standard of the experimental fish. After the gill cover stops flapping and there is no response to the needle, the experimental fish is placed in the natural water temperature and does not recover within 5 minutes. Activity, repeat the experiment 3 times, select the highest temperature when dead fish appear, as the lethal temperature of fish.

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