Method for simultaneously determining various flavor nucleotides in livestock and poultry meat products and application

Abstract

The invention provides a method for simultaneously determining various flavor nucleotides in a livestock and poultry meat product and application, and the method comprises the following steps: freeze-drying the livestock and poultry meat product, and then preparing the freeze-dried livestock and poultry meat product into a test solution; measuring the contents of CMP (cytidylic acid), ATP (adenosine triphosphate), GMP (guanine monophosphate), IMP (inosinic acid), ADP (adenosine diphosphate), Hy (hypoxanthine), AMP (adenosine monophosphate) and IN (inosine) in the test solution by adopting HPLC (high performance liquid chromatography); in HPLC, a mobile phase A is a phosphate buffer solution with the volume concentration of 95%, a mobile phase B is methanol with the volume concentration of 5%, gradient elution is carried out, and the gradient elution condition is 0-11 min, 99% phosphate buffer solution and 1% methanol; when 12 min, 80% phosphate buffer solution and 20% methanol are added, and the conditions are maintained for 6-7 min; the phosphate buffer solution is heated to 99% within 2 minutes, and then the temperature is kept for 8-10 minutes.

Description

Technical Field

[0001] The present invention relates to the technical field of food detection, and particularly relates to a method and application for simultaneously determining multiple flavor nucleotides in livestock and poultry meat products. Background Art

[0002] The flavor nucleotides in livestock and poultry meat are important flavor substances, mainly including 5'-inosinic acid (5'-IMP), 5'-guanylic acid (5'-GMP) and adenylic acid (AMP). The content of these substances significantly affects the flavor characteristics of livestock and poultry meat. Accurately evaluating the content of each flavor nucleotide in livestock and poultry meat is of great significance for regulating the flavor of livestock and poultry meat. At present, there is no national standard for the method of evaluating flavor nucleotides in livestock and poultry meat in the laboratory. There is only the group standard T / NAIA 003-2020 "Determination of inosine monophosphate in muscle - High performance liquid chromatography method". The group standard only has a method for determining inosinic acid, lacking evaluation methods and standards for other flavor substances. The above method for detecting inosinic acid in muscle uses fresh meat. On the one hand, since more than 70% of the meat sample is water and there is water loss during processing, it will cause a large deviation in the results and poor data stability. On the other hand, during the pre - treatment of the experiment, fresh meat is easily contaminated and bacteria are likely to breed, affecting the detection results. Summary of the Invention

[0003] The present invention aims to provide a method and application for simultaneously determining multiple flavor nucleotides in livestock and poultry meat products to fill the blank of simultaneous detection of multiple flavor nucleotides in livestock and poultry meat at present, and to solve the problem that the existing method for determining inosinic acid cannot accurately evaluate the flavor characteristics of livestock and poultry meat. The method of the present invention can simultaneously detect the content of at least three flavor nucleotides in livestock and poultry meat and reduce the influence of moisture on the detection process.

[0004] To achieve the above - mentioned technical purpose, the technical solution adopted by the present invention is as follows:

[0005] In the first aspect, the present invention provides a method for simultaneously determining multiple flavor nucleotides in livestock and poultry meat products, including: freeze - drying the livestock and poultry meat products, then preparing the freeze - dried livestock and poultry meat products into a test solution, and using HPLC to determine the contents of CMP (cytidine monophosphate), ATP (adenosine triphosphate), GMP (guanosine monophosphate), IMP (inosinic acid), ADP (adenosine diphosphate), Hy (hypoxanthine), AMP (adenylic acid), and IN (inosine) in the test solution;

[0006] In the HPLC, mobile phase A is a phosphate buffer solution with a volume concentration of 95%, and mobile phase B is methanol with a volume concentration of 5%. Gradient elution is performed, and the gradient elution conditions are as follows: from 0 to 11 min, 99% phosphate buffer solution and 1% methanol; at 12 min, 80% phosphate buffer solution and 20% methanol, and this is maintained for 6 - 7 min; within 2 minutes, the phosphate buffer solution is increased to 99%, and then maintained for 8 - 10 min.

[0007] Preferably, in the HPLC, the detection wavelength is 250 - 260 nm; the flow rate is 0.1 - 1.0 mL / min; the column temperature is 20 - 25 °C.

[0008] Preferably, in the HPLC, the detection wavelength is 254 nm; the flow rate is 0.6 mL / min; the column temperature is 25 °C.

[0009] Preferably, the preparation method of the test solution includes: adding a perchloric acid solution to the freeze - dried livestock and poultry meat products for ultrasonic extraction, then performing solid - liquid separation, adding the perchloric acid solution to the precipitate for repeated extraction several times, and combining the supernatants from multiple extractions and adjusting the pH value.

[0010] Preferably, in the preparation method of the test solution, the weight - to - volume ratio of the freeze - dried livestock and poultry meat products to the perchloric acid solution for two times is: 0.1 - 0.15:1:1; further preferably, the mass concentration of the perchloric acid solution is 3 - 7%, more preferably 5%; and / or, adding the perchloric acid solution to the precipitate for repeated extraction 1 time; and / or, combining the supernatants from multiple extractions and adjusting the pH value to 5.0 - 6.0, preferably 5.8.

[0011] Preferably, the perchloric acid solution for two times is pre - cooled at 0 - 4 °C for 1 - 2 h before use.

[0012] Preferably, the freeze - drying of the livestock and poultry meat products includes: first pre - freezing the livestock and poultry meat products, further preferably, the control parameters for pre - freezing are: - 70 - 80 °C, 2 - 5 h; then performing gradient freeze - drying on the pre - frozen livestock and poultry meat products, further preferably, the program for gradient freeze - drying is: under vacuum, - 55 - 35 °C, 4 - 6 h; - 35 - 15 °C, 2 - 4 h; - 15 - 5 °C, 5 - 8 h; 5 - 25 °C, 10 - 15 h; 25 - 30 °C, 1 - 3 h.

[0013] More preferably, in the freeze-drying process, the control parameters for pre-freezing are: -78 to -80°C, 2.5 to 3.5 h; and / or, the program for gradient freeze-drying is: under vacuum, -50 ± 1°C, 4 h ± 10 min; -40 ± 1°C, 1 h ± 10 min; -30 ± 1°C, 1 h ± 10 min; -20 ± 1°C, 2 h ± 10 min; -10 ± 1°C, 5 h ± 10 min; 0 ± 1°C, 2 h ± 10 min; 10 ± 1°C, 8 h ± 10 min; 20 ± 1°C, 6 h ± 10 min; 25 ± 1°C, 2 h ± 10 min.

[0014] Preferably, the livestock and poultry meat products are subjected to comminution treatment before pre-freezing.

[0015] In a second aspect, the present invention provides a method for evaluating the taste characteristics of livestock and poultry meat. The contents of CMP (cytidine monophosphate), ATP (adenosine triphosphate), GMP (guanosine monophosphate), IMP (inosine monophosphate), ADP (adenosine diphosphate), Hy (hypoxanthine), AMP (adenosine monophosphate), and IN (inosine) in the livestock and poultry meat samples are obtained by the method described in the first aspect, and the evaluation result of the taste characteristics is obtained based on the contents of each nucleotide substance.

[0016] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0017] The present invention explores a method for simultaneously determining multiple flavor nucleotides in livestock and poultry meat products, changing the previous method of directly using fresh meat to determine a single flavor nucleotide: inosine monophosphate. On the one hand, it avoids the large deviation in detection results caused by the loss of moisture in the meat; on the other hand, it reduces the reproduction of bacteria and the contamination of chicken caused by the long treatment time of fresh meat. In addition, the present invention can simultaneously determine the contents of 8 flavor nucleotides, namely CMP (cytidine monophosphate), ATP (adenosine triphosphate), GMP (guanosine monophosphate), IMP (inosine monophosphate), ADP (adenosine diphosphate), Hy (hypoxanthine), AMP (adenosine monophosphate), and IN (inosine) in one experiment, improving the detection efficiency and reducing the detection cost. Therefore, this method is convenient, fast, accurate, and can more accurately evaluate the taste characteristics of livestock and poultry meat. Description of the Drawings

[0018] Figure 1 : Elution profiles of multiple flavor nucleotides. Figure 1 This is the HPLC result of the chicken sample in Example 1 of the present invention. Detailed Embodiments

[0019] In the description of the present invention, it should be noted that for those not specified in the examples, they are carried out according to conventional conditions or the conditions recommended by the manufacturer. For the reagents or instruments not specified by the manufacturer, they are all conventional products that can be obtained through commercial purchase.

[0020] In a specific embodiment of the present invention, a method for simultaneously determining multiple flavor nucleotides in livestock and poultry meat products is provided. Taking chicken as an example, the method includes the following steps: freeze-drying the chicken product, and then preparing a test solution from the freeze-dried chicken product. HPLC is used to determine the contents of CMP (cytidine monophosphate), ATP (adenosine triphosphate), GMP (guanosine monophosphate), IMP (inosine monophosphate), ADP (adenosine diphosphate), Hy (hypoxanthine), AMP (adenylic acid), and IN (inosine) in the test solution; wherein, mobile phase A in HPLC is a phosphate buffer solution with a volume concentration of 95%, and mobile phase B is methanol with a volume concentration of 5%. Gradient elution is carried out, and the gradient elution conditions are as follows: 0 - 11 min, 99% phosphate buffer solution and 1% methanol; at 12 min, 80% phosphate buffer solution and 20% methanol, and maintain for 6 - 7 min; the phosphate buffer solution rises to 99% within 2 minutes, and then maintain for 8 - 10 min. Other detection conditions also include: the detection wavelength is 250 - 260 nm; the flow rate is 0.1 - 1.0 mL / min; the column temperature is 20 - 25 °C. Preferably, the detection wavelength is 254 nm; the flow rate is 0.6 mL / min; the column temperature is 25 °C.

[0021] The method also includes the preparation of the test solution: adding perchloric acid solution to the freeze-dried chicken product for ultrasonic extraction, then performing solid-liquid separation, adding the perchloric acid solution to the precipitate for repeated extraction several times, and combining the supernatants of multiple extractions and adjusting the pH value. Among them, the weight-volume ratio of the freeze-dried chicken product to the perchloric acid solution for two times is: 0.1 - 0.15:1:1; the preferred mass concentration of the perchloric acid solution is 3 - 7%, and more preferably 5%. In addition, repeating the extraction once by adding perchloric acid solution to the precipitate has a better effect, which can ensure that multiple flavor nucleotides are extracted into the solution. The supernatants of multiple extractions are combined and the pH value is adjusted to 5.8. In actual operation, the pH can be controlled between 5.0 and 6.0. If the pH is too low or too high, the structure of nucleotides will be damaged, resulting in large variations in the detection results. The perchloric acid solution is pre-cooled at 0 - 4 °C for 1 - 2 h before use. Pre-cooling the perchloric acid solution can reduce volatilization on the one hand and reduce the reaction rate on the other hand, thus avoiding the splashing of the test solution caused by the generation of a large amount of heat.

[0022] In a preferred embodiment of the present invention, the process of freeze-drying chicken products includes: first pre-freezing the chicken products, and the preferred control parameters for pre-freezing are: -70 to -80 °C for 2 to 5 hours; then subjecting the pre-frozen chicken products to gradient freeze-drying, and the preferred program for gradient freeze-drying is: under vacuum, -55 to -35 °C for 4 to 6 hours; -35 to -15 °C for 2 to 4 hours; -15 to 5 °C for 5 to 8 hours; 5 to 25 °C for 10 to 15 hours; 25 to 30 °C for 1 to 3 hours. This freeze-drying process can fully remove the moisture in the chicken without affecting the active ingredients of the chicken.

[0023] The most preferred control parameters for pre-freezing in freeze-drying are: -78 to -80 °C for 2.5 to 3.5 hours; the program for gradient freeze-drying is: under vacuum, -50 ± 1 °C for 4 hours ± 10 minutes; -40 ± 1 °C for 1 hour ± 10 minutes; -30 ± 1 °C for 1 hour ± 10 minutes; -20 ± 1 °C for 2 hours ± 10 minutes; -10 ± 1 °C for 5 hours ± 10 minutes; 0 ± 1 °C for 2 hours ± 10 minutes; 10 ± 1 °C for 8 hours ± 10 minutes; 20 ± 1 °C for 6 hours ± 10 minutes; 25 ± 1 °C for 2 hours ± 10 minutes. In addition, pulverizing the chicken products before pre-freezing can ensure the efficiency of freeze-drying.

[0024] The present invention will be further described in detail below in conjunction with the accompanying drawings and specific embodiments, which are explanations of the present invention rather than limitations.

[0025] Example 1

[0026] This example provides a method for simultaneously determining multiple flavor nucleotides in chicken products. The chicken products used are from the yellow-feathered broiler native 2 roosters in the broiler experimental farm of Wenshi Group. The specific method is as follows:

[0027] ① Process the meat sample. Use a meat grinder to evenly pulverize the meat. After pulverization, weigh a certain weight (m1) of fresh meat and put it into a weighed (m0) tin box.

[0028] ② Put the tin box together with the meat into a freezer at -80 °C and place it for 3 hours. At the same time, balance the cold trap temperature of the vacuum freeze-dryer to -50 °C, and then set the freeze-drying program for livestock and poultry meat (see Table 1 below).

[0029] Table 1 Chicken freeze-drying program

[0030] Stage Temperature (°C) Vacuum (Pa) Time (h) Remarks 1 -50 0 4 2 -40 0 1 Evacuation stage 3 -30 0 1 4 -20 0 2 5 -10 0 5 6 0 0 2 7 10 0 8 8 20 0 6 9 25 0 2 Constant temperature stage

[0031] ③ Take out the tin box containing meat frozen at -80 °C and put it into the previously prepared vacuum freeze-dryer.

[0032] ④ According to the program set by the vacuum freeze-dryer, subject the meat product to freeze-drying.

[0033] ⑤ Take out the freeze-dried meat sample, weigh it and record as m2, then seal it and store it in a -20°C refrigerator for future testing. At the same time, the moisture content a of the livestock and poultry meat can be known as:

[0034]

[0035] ⑥ Set the high performance liquid chromatography conditions:

[0036] Chromatographic column: TC-C18 column (250mm * 4.6um (i.d.), particle size 5μm);

[0037] Column temperature: 25°C;

[0038] Mobile phase: phosphate buffer solution + methanol (95 + 5);

[0039] Flow rate: 0.60 mL / min;

[0040] Injection volume: 20 μL;

[0041] Detection wavelength: 254 nm.

[0042] Gradient elution setting: 0 - 10 min, 99% phosphate buffer solution and 1% methanol; at 12 min, 80% phosphate buffer solution and 20% methanol are maintained for 6 min; within 2 min, the phosphate buffer solution is increased to 99% and then maintained for 8 min.

[0043] ⑦ Preparation of test solution

[0044] Weigh 2 - 3 g (accurate to 0.01 g, the chicken sample in this example is 2.00 g) of the minced and homogenized chicken sample into a 50 mL centrifuge tube, and add 20 mL of 5% perchloric acid solution pre-cooled in a 4°C refrigerator for 1 hour. Then extract ultrasonically for 5 min, centrifuge at 4000 rpm for 10 min at 4°C, and take the supernatant. Add another 20 mL of pre-cooled 5% perchloric acid solution to the precipitate. Repeat the above steps, combine all the supernatants in a 50 mL conical flask, and adjust the pH to 5.8 with 0.5 mol / L potassium hydroxide solution. Accurately pipette 20 mL from the adjusted pH extract and make up the volume to 100 mL in a volumetric flask. Take an appropriate amount of the made-up test solution, filter it through a 0.22 μm filter membrane into an injection vial for on-machine determination. The elution conditions have been tested, and the optimal determination results are as shown in Table 2 and Figure 1 shown.

[0045] Table 2 HPLC determination results of chicken samples

[0046]

[0047]

[0048] In addition, before adopting the elution conditions corresponding to the data in Table 2, the elution conditions adopted in the present invention were as follows: from 0 to 9 minutes, 99% phosphate buffer solution and 1% methanol; at 10 minutes, 80% phosphate buffer solution and 20% methanol, and maintained for 5 to 6 minutes; within 1 minute, the phosphate buffer solution was increased to 99%, and then maintained for 10 to 12 minutes. Other elution conditions were the same as the detection conditions described above. Finally, Table 3 below was obtained. From the results in Table 3, the flavor nucleotide substances in the chicken samples were not separated. Finally, the gradient elution program described above was further optimized (from 0 to 11 minutes, 99% phosphate buffer solution and 1% methanol; at 12 minutes, 80% phosphate buffer solution and 20% methanol, and maintained for 6 to 7 minutes; within 2 minutes, the phosphate buffer solution was increased to 99%, and then maintained for 8 to 10 minutes).

[0049] Table 3 Optimization of Elution Program

[0050]

[0051]

[0052] When determining the flavor nucleotides in the freeze-dried samples, the content Ci of 8 umami nucleotides including 5'-IMP, 5'-GMP, AMP, CMP, ATP, ADP, In, and Hy in the samples was expressed as mass fraction, with the unit of mg / g. It was calculated according to the formula Ci = (Cs * Ai * V) / (As * m). In the formula:

[0053] Ci—the content of various substances in the sample (mg / g);

[0054] Cs—the concentration of various substances in the standard working solution (mg / mL);

[0055] Ai—the peak area response value corresponding to various substances in the sample;

[0056] As—the peak area response value corresponding to various substances in the standard working solution;

[0057] V—the total volume of the sample extraction solution (mL);

[0058] m—the mass of the sample (g).

[0059] Based on the content of various flavor nucleotides in the freeze-dried chicken, through moisture conversion, the content of various substances in the fresh chicken can be obtained.

[0060] The method of the present invention is simple to operate, with less loss of nutrients in the meat samples, the detection process is simple, and the detection accuracy and efficiency are improved.

[0061] Example 2

[0062] Methodology Verification

[0063] (1) Standard curve

[0064] Prepare standard working solutions of eight substances, including standard working solutions containing 5'-IMP at 12.5, 25, 50, 62.5, 125 μg / mL, standard working solutions containing 5'-GMP, 5'-CMP, 5'-AMP, ATP, ADP, Hy at 2.5, 5, 10, 12.5, 25 μg / mL, and standard working solutions with a concentration gradient of In at 3.125, 6.25, 12.5, 31.25, 62.5 μg / mL. Perform chromatographic analysis according to the HPLC chromatographic conditions set in Example 1. Using the concentration of each substance as the independent variable (X) and the corresponding chromatographic peak area response value as the dependent variable (Y), establish the standard curve equation. The results show that: (1) In the concentration range of 12.5 - 125 μg / mL, there is a good linear relationship between the 5'-IMP concentration and the peak area response value. The regression equation is calculated as: Y = 36.007×X + 4.9022, and the correlation coefficient R 2 = 0.9998. (2) In the concentration range of 2.5 - 25 μg / mL, there is a good linear relationship between the 5'-GMP concentration and the peak area response value. The regression equation is calculated as: Y = 59.225×X - 4.3408, and the correlation coefficient R 2 = 0.9999; there is a good linear relationship between the 5'-CMP concentration and the peak area response value. The regression equation is calculated as: Y = 37.086×X + 7.11153, and the correlation coefficient R 2 = 0.9998; there is a good linear relationship between the 5'-AMP concentration and the peak area response value. The regression equation is calculated as: Y = 69.853×X + 302.53, R 2 = 0.9998; there is a good linear relationship between the 5'-ATP concentration and the peak area response value. The regression equation is calculated as: Y = 44.341×X - 17.824, R 2 = 0.9998; there is a good linear relationship between the 5'-ADP concentration and the peak area response value. The regression equation is calculated as: Y = 56.039×X - 16.645; R 2 = 0.9997; there is a good linear relationship between the Hy concentration and the peak area response value. The regression equation is calculated as: Y = 145.74×X + 7.6301;

[0065] R 2 = 0.9996; (3) In the concentration range of 2.5 - 25 μg / mL, there is a good linear relationship between the AMP concentration and the peak area response value. The regression equation is calculated as: Y = 69.853×X + 302.53, and the correlation coefficient R 2= 0.9998. The standard curve data of eight nucleotide substances are shown in Table 4.

[0066] Table 4 Standard Curve Data of Eight Nucleotide Substances

[0067]

[0068]

[0069] (2) Recovery Rate Determination

[0070] Prepare the sample solution according to the method provided in ⑦ of Example 1. Take one portion and transfer 20 mL of the test solution directly to a 100 mL volumetric flask and make up the volume to the mark with water. For the other portion, transfer 20 mL of the test solution, then add 1 mL of 1.00 mg / mL 5'-IMP, 1 mL of 200 μg / mL 5'-GMP, 1 mL of 100 μg / mL AMP, 1 mL of 100 μg / mL CMP, 1 mL of 200 μg / mL ATP, 2 mL of 100 μg / mL ADP, 1 mL of 200 μg / mL In, and 1 mL of 200 μg / mL Hy, and then make up the volume to the mark with water in a 100 mL volumetric flask, and then conduct the recovery test. The recovery rate data of 8 substances are shown in Table 5.

[0071] Table 5 Results of Recovery Tests for 8 Substances

[0072]

[0073]

[0074] According to the data in Table 5, it can be seen that the recovery rates of the 8 components all reach over 90%, and the recovery rates are relatively high.

[0075] (3) Precision Determination

[0076] According to the method established in this experiment, repeat the detection 5 times. Calculate the average value and relative standard deviation (RSD) from the repeated measurement data. It is required that the absolute difference between any two of the 5 independent measurement results does not exceed 10% of the arithmetic mean of these two measurement values to meet the test requirements. The results are shown in Table 6.

[0077] Table 6 Results of Precision Tests for 8 Substances

[0078]

[0079]

[0080] According to the data in Table 6, the RSD values in the precision determination results of the 8 substances are lower than 2%, and the effect is good.

[0081] Example 3

[0082] This embodiment provides a method for evaluating the taste characteristics of chicken. The chicken samples used are from the yellow - feather broiler native 2 roosters in the broiler test farm of Wenshi Group, which are not the same batch as the samples in Example 1. Using the method of Example 1, the contents of CMP (cytidine monophosphate), ATP (adenosine triphosphate), GMP (guanosine monophosphate), IMP (inosine monophosphate), ADP (adenosine diphosphate), Hy (hypoxanthine), AMP (adenosine monophosphate), and IN (inosine) in the livestock and poultry meat samples are 0.08 mg / kg, 0.13 mg / kg, 0.12 mg / kg, 4.21 mg / kg, 0.26 mg / kg, 0.15 mg / kg, 0.12 mg / kg, and 0.54 mg / kg respectively. It can be seen from the test results that the content of IMP (inosine monophosphate), which represents the umami taste of this chicken sample, is significantly higher than that of other nucleotide substances. Based on this result, it can be known that the taste characteristics of this sample are better.

[0083] In summary, through the freeze - drying treatment of fresh meat samples, the present invention protects the nutrients in the meat products and reduces the loss of meat samples. Multiple substances can be detected simultaneously in one experiment, which improves the detection efficiency while ensuring the accuracy of the results and the timeliness and convenience of detection. In addition, the present invention can also reduce environmental pollution, reduce the spoilage of meat samples and the loss of nutrients caused by long - term storage of meat samples, and at the same time reduce the waste of substances and environmental pollution caused by the deterioration of meat samples. The method of this invention is simple to operate, with less loss of nutrients in meat samples, and the detection process is simple, which improves the detection accuracy and efficiency.

[0084] The above - described embodiments only represent several implementation manners of the present invention. Their descriptions are relatively specific and detailed, but they should not be construed as limiting the scope of the patent of the present invention. It should be noted that for those of ordinary skill in the art, without departing from the concept of the present invention, several modifications and improvements can still be made, and these all belong to the protection scope of the present invention. Therefore, the protection scope of the patent of the present invention should be subject to the appended claims.

Claims

1. Method for simultaneously determining multiple flavor nucleotides in livestock and poultry meat products, characterized in that, Comprising: Freeze-drying livestock and poultry meat products, then formulating the freeze-dried livestock and poultry meat products into a test solution, and measuring the contents of CMP (cytidine monophosphate), ATP (adenosine triphosphate), GMP (guanosine monophosphate), IMP (inosine monophosphate), ADP (adenosine diphosphate), Hy (hypoxanthine), AMP (adenylic acid), and IN (inosine) in the test solution by HPLC; In the HPLC, mobile phase A is a phosphate buffer solution with a volume concentration of 95%, and mobile phase B is methanol with a volume concentration of 5%. Gradient elution is performed, and the gradient elution conditions are as follows: 0 - 11 min, 99% phosphate buffer solution and 1% methanol; at 12 min, 80% phosphate buffer solution and 20% methanol, and maintain for 6 - 7 min; the phosphate buffer solution rises to 99% within 2 minutes, and then maintain for 8 - 10 min.

2. The method according to claim 1, wherein In the HPLC, the detection wavelength is 250 - 260 nm; the flow rate is 0.1 - 1.0 mL / min; the column temperature is 20 - 25 °C.

3. The method according to claim 1 or 2, characterized in that, In the HPLC, the detection wavelength is 254 nm; the flow rate is 0.6 ml / min; the column temperature is 25 °C.

4. The method according to any one of claims 1 to 3, characterized in that The method for formulating the test solution includes: adding a perchloric acid solution to the freeze-dried livestock and poultry meat products for ultrasonic extraction, then performing solid-liquid separation, adding the perchloric acid solution to the precipitate for repeated extraction several times, and combining the supernatants obtained from multiple extractions and adjusting the pH value.

5. The method according to claim 4, wherein In the method for formulating the test solution, the weight-volume ratio of the freeze-dried livestock and poultry meat products to the perchloric acid solution twice is: 0.1 - 0.15:1:1; preferably, the mass concentration of the perchloric acid solution is 3 - 7%, more preferably 5%; and / or, adding the perchloric acid solution to the precipitate for repeated extraction 1 time; and / or, combining the supernatants obtained from multiple extractions and adjusting the pH value to 5.0 - 6.0, preferably 5.

8.

6. The method according to claim 5, wherein The perchloric acid solution twice is pre-cooled at 0 - 4 °C for 1 - 2 h before use.

7. The method according to any one of claims 1 to 6, characterized in that The freeze-drying of the livestock and poultry meat products includes: pre-freezing the livestock and poultry meat products first. Preferably, the control parameters for pre-freezing are: -70 - -80 °C, 2 - 5 h; then performing gradient freeze-drying on the pre-frozen livestock and poultry meat products. Preferably, the program for gradient freeze-drying is: under vacuum, -55 - -35 °C, 4 - 6 h; -35 - -15 °C, 2 - 4 h; -15 - 5 °C, 5 - 8 h; 5 - 25 °C, 10 - 15 h; 25 - 30 °C, 1 - 3 h.

8. The method according to claim 7, wherein In the freeze-drying, the control parameters for pre-freezing are: -78 - -80 °C, 2.5 - 3.5 h; and / or, the program for gradient freeze-drying is: under vacuum, -50 ± 1 °C, 4 h ± 10 min; -40 ± 1 °C, 1 h ± 10 min; -30 ± 1 °C, 1 h ± 10 min; -20 ± 1 °C, 2 h ± 10 min; -10 ± 1 °C, 5 h ± 10 min; 0 ± 1 °C, 2 h ± 10 min; 10 ± 1 °C, 8 h ± 10 min; 20 ± 1 °C, 6 h ± 10 min; 25 ± 1 °C, 2 h ± 10 min.

9. The method according to claim 7 or 8, characterized in that The livestock and poultry meat products are pulverized before pre-freezing.

10. A method for evaluating the taste characteristics of livestock and poultry meat, characterized in that, The contents of CMP (cytidine monophosphate), ATP (adenosine triphosphate), GMP (guanosine monophosphate), IMP (inosine monophosphate), ADP (adenosine diphosphate), Hy (hypoxanthine), AMP (adenylic acid), and IN (inosine) in the livestock and poultry meat samples are obtained by using the method according to any one of claims 1 to 9, and the evaluation result of the taste characteristics is obtained based on the contents of each nucleotide substance.

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