A method for evaluating freshness quality of ice-freshed large yellow croaker by using bioimpedance technology

Abstract

The present application relates to the field of non-destructive detection of fish freshness, and particularly relates to a method for evaluating the freshness quality of ice-fresh big yellow croaker by using bioimpedance technology. The method for evaluating the freshness quality of ice-fresh big yellow croaker by using bioimpedance technology comprises the following steps: S1, immediately killing and processing fresh and healthy farmed big yellow croaker after being captured, and then storing the big yellow croaker in ice; S2, measuring the back and abdominal muscle bioimpedance values of the ice-fresh big yellow croaker in S1 at a frequency of 2 kHz or 100 kHz by using an impedance analyzer; S3, substituting the bioimpedance values Z 2kHz or Z 100 kHz measured in S2 into a big yellow croaker impedance and each quality change prediction model to calculate the values of key components and freshness indexes, wherein the key components include protein, fat and moisture, and the freshness indexes include color difference, pH, water activity, juice loss rate and total bacterial count; and S4, directly using the calculated values to replace the measured values to evaluate the fish body quality.

Description

Technical Field

[0001] This invention relates to the field of non-destructive testing of fish freshness, and in particular to a method for evaluating the freshness and quality of chilled large yellow croaker using bioimpedance technology. Background Technology

[0002] Large yellow croaker, also known as yellow croaker, belongs to the order Perciformes, family Sciaenidae, and genus Croaker. It is a species endemic to my country, widely distributed, with Zhejiang and Fujian being the main aquaculture areas. Its attractive golden color, tender and delicious flesh, and high content of protein and polyunsaturated fatty acids have earned it the reputation of "national fish of the sea," making it a popular choice among consumers and an important economic fish species in my country's marine aquaculture industry. With the upgrading of consumption among urban and rural residents, the demand for large yellow croaker is increasing, and higher requirements are being placed on its freshness and quality.

[0003] Because large yellow croaker dies immediately after being caught, it is currently mainly sold in the market as chilled fish. The freshness and safety of chilled large yellow croaker directly affect its sales value, processing value, and food safety. Therefore, rapid and effective monitoring of changes in the freshness and quality of fresh large yellow croaker is of great significance. During storage, chilling cannot inhibit the activity of endogenous proteases and the growth and reproduction of microorganisms in large yellow croaker, leading to physical, chemical, and biological changes. This results in varying degrees of loss of nutrients in the fish meat, a decline in the freshness and quality of the fish, and impacts production and sales. Large yellow croaker of different freshness levels can be processed into different products; therefore, there is an urgent need to establish a method to quickly predict changes in the freshness and quality of chilled large yellow croaker, thereby reducing economic losses in commercial activities.

[0004] Traditional indicators such as protein, fat, moisture, color difference, pH, water activity, juice loss rate, and total bacterial count are commonly used as important indicators of changes in fish freshness and quality, accurately reflecting changes in the freshness and spoilage of fish meat. After fish die, during rigor mortis, glycogen degradation in the muscle produces lactic acid, causing a decrease in pH. Subsequently, under the action of enzymes and microorganisms, amino acids and proteins degrade, producing alkaline substances such as ammonia and amines, causing the pH to rise. During refrigeration, fish undergo protein denaturation, fat oxidation, cell dehydration, gradual juice loss, changes in fish meat color, and an increase in microbial numbers, leading to a decline in fish quality. Currently, traditional methods for testing the freshness and quality of aquatic products are generally destructive, time-consuming, labor-intensive, material-intensive, and cumbersome, failing to meet the current needs of enterprises, supermarkets, market regulators, and consumers for rapid, non-destructive, and accurate testing.

[0005] In recent years, novel detection methods have gradually become a research and application hotspot, such as odor fingerprinting, sensory bionics, spectroscopy, and biosensors. These rapid detection methods offer advantages such as speed, non-destructive testing, and wide applicability, but their high cost and portability hinder their widespread adoption. Bioimpedance technology is a non-destructive testing method that utilizes the electrical properties and changes in an organism to extract tissue state information. It typically uses an electrode system placed on the body surface to deliver a small alternating current or voltage to the object being tested, obtaining electrical information reflecting the relevant physicochemical state. It is non-destructive, rapid, portable, inexpensive, and provides rich information, showing great potential for application in the field of non-destructive testing of fish. Currently, there is no known method for non-destructive monitoring and evaluation of fish quality based on the bioimpedance of large yellow croaker during ice storage. Summary of the Invention

[0006] The purpose of this invention is to provide a method for evaluating the freshness and quality of chilled large yellow croaker using bioimpedance technology. This method utilizes bioimpedance technology to quickly and non-destructively detect the overall condition of the fish, overcoming the problems of time-consuming, cumbersome, and destructive nature of traditional freshness index determination.

[0007] The technical solution adopted by this invention to solve its technical problem is:

[0008] A method for assessing the freshness and quality of chilled large yellow croaker using bioimpedance technology, the method comprising the following steps:

[0009] S1. Freshly caught, healthy farmed large yellow croakers are immediately killed, processed, and then frozen.

[0010] S2. Use an impedance analyzer to measure the bioimpedance values ​​of the dorsal and abdominal muscles of the fresh yellow croaker in S1 at frequencies of 2kHz or 100kHz.

[0011] S3, The bioimpedance value Z measured in S2 2kHz or Z 100 kHz Substitute the values ​​into the prediction model for impedance and quality variations of large yellow croaker.

[0012]

[0013]

[0014] Among them, Z 2kHz Z represents the fish body impedance value measured at a current frequency of 2kHz. 100kHz The fish impedance value was measured at a current frequency of 100kHz.

[0015] The values ​​of key components and freshness indicators were calculated. Key components include protein, fat, and moisture. Freshness indicators include color difference, pH, water activity, juice loss rate, and total bacterial count.

[0016] S4. Directly use the calculated values ​​instead of the measured values ​​to evaluate the quality of the fish.

[0017] The physicochemical manifestations of fish after death include pH fluctuations and ion translocation (Ca). 2+ K + Changes in the physicochemical properties of fish, such as protein degradation and lipid oxidation, manifest structurally as alterations in myofibrils, intracellular and extracellular fluid spaces, cell membrane channels, phospholipid bilayers, and extracellular matrix. These changes significantly alter the electrical properties of the fish. Based on the changes in key components (protein, fat, and moisture) and freshness indicators (color difference, pH, water activity, juice loss rate, and total bacterial count) of large yellow croaker stored at 4°C, this invention establishes a predictive model for the changes in impedance and freshness quality of large yellow croaker. This model allows for a more intuitive, accurate, and comprehensive assessment of the overall changes within the fish.

[0018] In the evaluation method of this invention, each quality indicator has a prediction function model. During the test, the prediction function model is called according to the actual needs, and the freshness of the fish is judged based on the value of the component or freshness indicator, so as to achieve rapid and non-destructive quality detection.

[0019] Because the impedance of large yellow croaker peaks during the initial stage of ice storage (0-24h), the inventors used an impedance measurement model to assess fish quality in their research. They determined whether the measured impedance value was before or after the peak by measuring the fish's impedance twice, either during the ice storage period or at 15-minute intervals. 2kHz A significant increase indicates the pre-peak period, while no change or a slight decrease indicates the post-peak period.

[0020] To improve the accuracy and practicality of the fitting equation between impedance, storage time, and quality indicators, Y=Z is used under various frequency conditions. 24 h Using the horizontal line (peak impedance value at 24h) as the baseline, adjust the 0h impedance value (pre-peak data) vertically above the peak horizontal baseline (see hollow icon). Figure 2 Data analysis was conducted using both the pre-peak adjusted impedance value (hollow icon) and the post-peak measured impedance value (solid icon). Fitting the adjusted data revealed (red dashed line, see...) Figure 2The impedance values ​​at different frequencies showed a very strong functional relationship with the ice storage time (r = 0.9829–0.9965, P < 0.05). Generally, the impedance value of fish reaches its peak after 24 hours of storage, but current studies cannot utilize the 24-hour impedance value for quality analysis. This invention uses mathematical analysis to apply the 24-hour data to the model establishment, thereby improving the accuracy and convenience of the detection method.

[0021] Therefore, the present invention has the following beneficial effects:

[0022] 1. Using bioimpedance technology, the overall condition of fish can be detected quickly and non-destructively, overcoming the problems of time-consuming, cumbersome, and destructive traditional freshness index determination.

[0023] 2. It integrates the correlation between bioimpedance and various quality indicators such as protein, fat, moisture, color difference, pH, water activity, juice loss rate and total bacterial count, and is not limited to a single physicochemical or structural indicator. It can more comprehensively and accurately assess the overall changes of large yellow croaker during the freezing process.

[0024] 3. It can effectively promote the safety monitoring of aquatic products, optimize the allocation of fishery resources, and improve the optimal utilization level of aquatic products, thus effectively promoting the healthy development of the marine fishery economy and the aquatic product industry. Attached Figure Description

[0025] Figure 1 This describes the changes in the resistance values ​​of the dorsal and abdominal muscles of large yellow croaker during ice storage.

[0026] Figure 2 This describes the changes in protein, fat, and water content in the back and abdominal muscles of large yellow croaker during the freezing process.

[0027] Figure 3 This study investigated the changes in color difference, pH, water activity, juice loss rate, and total bacterial count of the back muscle of large yellow croaker during ice storage.

[0028] Figure 4 The relationship between predicted values ​​calculated using impedance function equations and predicted values ​​from direct chemical analysis for the dorsal and abdominal muscle components of frozen large yellow croaker;

[0029] Figure 5 The relationship between the predicted values ​​of dorsal muscle freshness of frozen large yellow croaker calculated using impedance function equations and the predicted values ​​of direct chemical analysis. Detailed Implementation

[0030] The technical solution of the present invention will be further described in detail below through specific embodiments. It should be understood that the implementation of the present invention is not limited to the following embodiments, and any modifications and / or alterations made to the present invention will fall within the protection scope of the present invention.

[0031] In this invention, unless otherwise specified, all parts and percentages are by weight, and the equipment and raw materials used are commercially available or commonly used in the art. Unless otherwise specified, the methods in the following embodiments are conventional methods in the art.

[0032] Example:

[0033] 1. Materials and Methods

[0034] 1.1 Raw Materials and Instruments

[0035] Raw materials: Large yellow croakers were purchased from Zhoushan Peninsula Aquaculture Co., Ltd., all of which are deep-sea cage-cultured fish from the Zhoushan waters of Zhejiang, China. The selection criteria were similar fatness and leanness, intact scales, strong vitality, and no disease. The large yellow croakers were immediately killed, processed, rinsed, and the muscle parts were removed and placed in sterile retort bags, placed in crushed ice, and stored in a refrigerator at 4°C.

[0036] Instruments: Fish Analyzer impedance analyzer (Yamato Corporation, Japan); CR-10 colorimeter (Konica Minolta); 723PC visible light spectrophotometer (Shanghai Jinghua Technology Instrument Co., Ltd.); 8695 pen pH meter (Hengxin Technology Co., Ltd.); LDZF-75L-I autoclave (Shanghai Shenan Medical Instrument Factory); SZT(F)-06A fat analyzer (Suzhou Tianwei Instrument Co., Ltd.); JC-HD intelligent water activity meter (Qingdao Juchuang Environmental Protection Group Co., Ltd.), etc.

[0037] 1.2 Test Methods

[0038] 1.2.1 Bioimpedance

[0039] The impedance of the dorsal and abdominal muscles of frozen large yellow croaker was measured at different frequencies (2, 5, 20, 50, 100 kHz) using a Fish Analyzer impedance analyzer.

[0040] 1.2.2 Protein

[0041] The total protein (TP) assay kit manufactured by Nanjing Jiancheng Biotechnology Co., Ltd. was used to determine the total protein content of the dorsal and abdominal muscles of frozen large yellow croaker. The specific operation was performed according to the kit instructions.

[0042] 1.2.3 Fat

[0043] 5.000g of back and abdominal muscle of frozen large yellow croaker were weighed separately, and the crude fat content of the back and abdominal muscle of frozen large yellow croaker was determined by Soxhlet extraction method according to GB 5009.6-2016 "National Food Safety Standard Determination of Fat in Food".

[0044] 1.2.4 Moisture

[0045] The moisture content of the dorsal and ventral muscles of frozen large yellow croaker was determined according to the direct drying method in GB 5009.3-2016 "National Food Safety Standard - Determination of Moisture in Food". 2.000g of the chopped sample was weighed, placed in a weighing bottle after constant weight determination, and dried in a 105℃ drying oven until constant weight was achieved.

[0046] 1.2.5 Water activity

[0047] The water activity was determined using the diffusion method with a water activity meter, as specified in GB 5009.238-2016, "National Food Safety Standard - Determination of Water Activity in Food". 2.000g of chopped back muscle from a large yellow croaker was weighed, quickly placed into a sample dish, the measuring chamber was sealed, and the measurement was performed.

[0048] 1.2.6 Color difference

[0049] Absorb excess moisture from the muscle surface with absorbent paper, and then measure the brightness (L) of the back muscles after calibration with a colorimeter. * ) and yellowness (b * ) Take measurements and record them (each measurement is taken at the location of the white flesh where there is no skin).

[0050] 1.2.7 pH

[0051] Place the calibrated pen pH meter on the surface of the dorsal muscle of a skinned fish and measure at different points on the same sample, recording all measurement points and their respective pH values.

[0052] 1.2.8 Juice loss rate

[0053] After the sample is removed from the refrigerator, weigh it (m1 / g), wipe off the juice on the sample surface with filter paper, weigh it again (m2 / g), and calculate the juice loss rate of the back and abdominal muscles according to the following formula.

[0054]

[0055] 1.2.9 Total bacterial count

[0056] The total bacterial count in the back muscles was determined according to the method in GB 4789.2-2016, "National Food Safety Standard - Microbiological Examination of Food - Determination of Total Colony Count".

[0057] 1.2.10 Impedance Model Accuracy Verification Experiment

[0058] By measuring the impedance values ​​of the dorsal and abdominal muscles of fish during ice storage at current frequencies of 2, 5, 20, 50, and 100 kHz, the predicted quality values ​​of large yellow croaker were calculated and substituted into a quality prediction model. Subsequently, physicochemical tests were performed on the dorsal and abdominal muscles of the fish using traditional quality indicator detection methods to obtain measured quality values. Linear regression analysis was used to calculate the correlation coefficient (r) between the predicted quality indicators and the measured chemical analysis values, thus verifying the practical application significance of the impedance prediction model.

[0059] 1.2.11 Data Processing and Analysis

[0060] Experimental data were processed using WPS Office software, and results are expressed as mean ± standard deviation. One-way ANOVA was performed using SPSS 26.0 software, and significance analysis was conducted using the Duncan test. P < 0.05 indicated significant differences, and P < 0.001 indicated highly significant differences. We analyzed the functional relationships between impedance at five current frequencies and components and freshness, and used the r-test to test their significance. The equation with the highest good fit and practical significance was selected as the ideal model for evaluating fish quality. Graphpad Prism 9 software was used for graphing.

[0061] 2 Results and Analysis

[0062] 2.1 Impedance Analysis

[0063] Resistance changes in frozen large yellow croaker, such as Figure 1 As shown, the impedance of the dorsal and abdominal muscles of the large yellow croaker exhibited a trend of first rapidly increasing and then slowly decreasing at various frequencies, reaching a peak after 24 hours of refrigeration. Subsequently, the impedance decreased at different rates until the end of storage. During refrigeration, the impedance of the dorsal muscles was higher than that of the abdominal muscles at different frequencies, and the impedance values ​​at lower frequencies were higher than those at higher frequencies. Refrigeration time affects impedance changes, and as the fish's muscle tissue stiffens, autolyzes, and decomposes, the cellular life state continuously declines, causing the difference in bioimpedance between high and low frequencies in the later stages of muscle tissue development to become increasingly smaller.

[0064] 2.2 Protein Analysis

[0065] Since changes in fish freshness and quality decline are primarily caused by changes in protein, protein degradation products may serve as potential quality assessment indicators or freshness indicators for aquatic products. For example... Figure 2 As shown in a, after the death of large yellow croaker, the protein content of the back and abdominal muscles decreased continuously with the extension of storage time. This may be due to the alteration and degradation of muscle protein molecular structure under the action of endogenous proteases in the muscle tissue and exogenous proteases from putrefactive bacteria.

[0066] 2.3 Fat Analysis

[0067] The changes in fat content of large yellow croaker under different freezing times are as follows: Figure 2 As shown in b, the fat content decreases continuously with the extension of storage time, indicating that the fat is continuously oxidized and decomposed.

[0068] 2.4 Moisture Analysis

[0069] The water-retention capacity of fish muscle is an important indicator of muscle quality, directly affecting the texture, freshness, and sensory quality of the fish meat. For example... Figure 2 As shown in c, the dorsal and abdominal muscles of large yellow croaker decreased with the extension of storage time under different ice storage times. The concentrations of intracellular and extracellular fluid in myofibril cells changed continuously, and water was transferred between the inside and outside of the cells, resulting in changes in the fish's impedance.

[0070] 2.5 Color Difference Analysis

[0071] The color of large yellow croaker is a decisive factor influencing consumer purchasing behavior and also an important factor affecting the shelf life of fresh fish. Table 1 shows the changes in color variation of large yellow croaker during refrigeration.

[0072] Table 1

[0073] Note: Different lowercase letters in different groups indicate significant differences (P < 0.05).

[0074] As shown in Table 1, over time, the L-shaped muscle of the large yellow croaker's back muscle... * The values ​​all showed a significant downward trend, decreasing from 47.89 (0h) to 44.07 (144h), indicating a decrease in the b value of the dorsal muscle of the large yellow croaker. * The value increased from 0.51 (0h) to 3.73 (144h), L * The continuously decreasing value indicates that the fish has lost juices during storage, resulting in a decrease in surface free water content and gloss. * The increase in values ​​may be related to increased lipid oxidation.

[0075] 2.6 pH Analysis

[0076] The pH of the muscle tissue of chilled large yellow croaker was measured from 0 h to 144 h of refrigeration, and the results are as follows: Figure 3 As shown in Figure a, the dorsal muscles of the large yellow croaker exhibit a "V-shaped" trend of first decreasing and then increasing throughout the storage process. The decrease in pH may be attributed to the formation of lactic acid, while the increase may be due to the increased production of volatile bases (e.g., ammonia, trimethylamine, histamine, etc.) generated by the decomposition of proteins, amino acids, and nitrogenous substances in the fish body under the action of endogenous or microbial enzymes.

[0077] 2.7 Water activity analysis

[0078] Water activity (Aw The equilibrium vapor pressure (EPP) of a food product is defined as the ratio of its equilibrium vapor pressure to the saturated vapor pressure of pure water at the same temperature. Its magnitude significantly influences the stability of a food's color, odor, and textural properties, and it has become one of the important indicators characterizing food stability and microbiological safety. Figure 3 b indicates that the initial water activity of the dorsal muscle of the large yellow croaker was 0.861. It increased the fastest after 24 hours of refrigeration, reaching 0.931, indicating a favorable environment for microbial reproduction. Subsequently, the water activity level tended to level off.

[0079] 2.8 Juice Loss Rate Analysis

[0080] Juice loss rate is a standard for measuring the water-holding capacity of fish meat. During freezing, large molecules such as proteins degrade, leading to water loss from the muscle and a continuous decrease in its water-holding capacity. Figure 3 As shown in c, during the refrigeration process, the loss rate of juice from the large yellow croaker showed an increasing trend, while the loss rate was relatively stable.

[0081] 2.9 Total Colony Count Analysis

[0082] Microorganisms are an important indicator for evaluating the freshness and safety of fish. Studies have shown that a total bacterial count of 6.001 g (CFU / g) or higher in large yellow croaker exceeds the safe consumption limit. The changes in total bacterial count of large yellow croaker at different storage times are shown below. Figure 3 As shown in d, by Figure 3 As can be seen from d, the number of microorganisms in the large yellow croaker showed an upward trend, with an initial total colony count of 3.071g (CFU / g), which reached the threshold at 96h.

[0083] 3. Impedance prediction model establishment and verification

[0084] 3.1 Establishment of the prediction model

[0085] We used power function analysis to examine the functional regression correlation between the 2, 5, 20, 50, and 100 kHz impedance values ​​(0-24h adjustment values, 48-144h measured values) of the dorsal and abdominal muscles of large yellow croaker at different freezing times and their components and freshness indicators. The functions with the strongest correlations are shown in Table 2. In actual testing, since the impedance of large yellow croaker peaks in the early stages of freezing, when using the impedance measurement model to assess fish quality, it is necessary to determine whether the measured impedance value is before or after the peak. Therefore, the tester needs to know the freezing time of the fish or measure the fish impedance twice at 15-minute intervals (Z2). kHz A significant increase indicates the pre-peak phase, while no change or a slight decrease indicates the post-peak phase. In actual testing of the quality of frozen large yellow croaker, the tester can measure the impedance values ​​of the back and abdominal muscles at different frequencies (2, 5, 20, 50, and 100 kHz) and use the established functional relationship equations between specific frequency impedance and components / freshness to assess the fish quality.

[0086] Table 2

[0087]

[0088] The measured fish body impedance value Z2 kHz or Z 100kHz By substituting these values ​​into the various prediction models in Table 2, the values ​​of various quality indicators of the fish can be obtained, thereby determining the quality of the fish. This method replaces the traditional experimental method for determining fish quality, saving fish resources, time, and laboratory reagents and consumables.

[0089] 3.2 Impedance Model Verification

[0090] Based on the functional equation model in Table 2, the muscle components and freshness indicators of another batch of frozen large yellow croaker were predicted, and the relationship between the model predictions and experimental measurements was evaluated using the Y=X equation. Figure 4 , 5 It was found that there was a significant correlation between the predicted and measured values ​​of muscle components and freshness of large yellow croaker. The measured values ​​were all obtained using traditional detection methods. Therefore, the validation demonstrates that the predictive model can be accurately used to assess the quality of chilled large yellow croaker.

[0091] 4. Summary

[0092] Experimental results showed that with prolonged storage time, the muscle impedance and various freshness quality indicators of large yellow croaker changed to varying degrees, and were significantly correlated (P < 0.05). Bioimpedance can reflect the changes in freshness of large yellow croaker during ice storage and can be used as a non-destructive and rapid method to assess the quality of large yellow croaker.

[0093] The various embodiments in this specification are described in a progressive manner, with each embodiment focusing on its differences from other embodiments. Similar or identical parts between embodiments can be referred to interchangeably. For the apparatus disclosed in the embodiments, since it corresponds to the method disclosed in the embodiments, the description is relatively simple; relevant parts can be referred to in the method section.

[0094] The above provides a detailed description of a method for evaluating the freshness and quality of chilled large yellow croaker using bioimpedance technology, as provided by this invention. Specific examples have been used to illustrate the principles and implementation methods of this invention. The descriptions of these embodiments are merely for the purpose of helping to understand the method and its core ideas. It should be noted that those skilled in the art can make various improvements and modifications to this invention without departing from its principles, and these improvements and modifications also fall within the protection scope of the claims of this invention.

Claims

1. A method for evaluating the freshness quality of ice-stored large yellow croaker by using bioimpedance technology, characterized in that The method comprises the following steps: S1, fresh captured healthy cultured large yellow croaker is stored at 4℃ after death; S2, the back and abdominal muscle bioimpedance values of the ice fresh large yellow croaker of S1 at 2 kHz or 100 kHz frequency are measured by using an impedance analyzer; S3, the bioimpedance value Z determined in S2 2kHz or Z 100 kHz Substitute the impedance of P. tridacnontus into the prediction model of each quality change, ， wherein Z 2kHz is the impedance value of the fish body measured at a current frequency of 2 kHz, Z 100kHz is the impedance value of the fish body measured at a current frequency of 100 kHz; The values of key components and freshness indicators are calculated, the key components including protein, fat and moisture, and the freshness indicators including brightness and yellowness, pH, water activity, juice loss rate and total bacterial count; When the impedance prediction model is used to evaluate the fish quality, it is necessary to judge whether the measured impedance value is before or after the peak; The measured impedance value is judged to be before or after the peak by measuring the impedance of the fish twice at an interval of 15 minutes or by the ice storage time of the fish; the impedance value is obviously increased before the peak, and there is no change or slight decrease after the peak; S4, the calculated values are used to evaluate the fish quality; The impedance of the large yellow croaker and each quality change prediction model is established under the condition of each frequency, and the horizontal line of Y=Z 24 h is the peak value level baseline. The data before the peak is adjusted vertically and equally above the peak value level baseline, and the data analysis is performed by jointly using the adjusted impedance value before the peak and the measured impedance value after the peak, wherein Z 24 h is the peak impedance value of 24 h.

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