Method for identifying grassland grazing sheep and grassland grazing mutton and application thereof

Abstract

The invention provides a grassland grazing sheep and grassland grazing mutton identification method and application thereof, and belongs to the technical field of food authenticity detection. The stable isotope values of delta 13C and delta 15N of the sheep neck hair are detected, and whether the sheep belongs to grassland grassland can be judged. The identification method for grassland grazing mutton provided by the invention comprises the following steps: detecting stable isotope values of delta 13C and delta 15N and the contents of fatty acid, flavor amino acid and total amino acid in mutton; if delta13C is less than or equal to-19.0 per mill, delta15N is more than or equal to 4.0 per mill, omega-6: omega-3 in fatty acid is less than or equal to 2.5, and flavor amino acid: total amino acid is more than or equal to 50%, determining that the mutton is grassland grazing mutton. According to the identification method, the essential characteristics of grassland grazing mutton are highlighted from a multi-index combined angle, the influence of factors such as variety, region, month age and man-made interference can be eliminated, and the identification precision reaches 100%.

Description

[0001] The present application is a divisional application of a patent application with the application date of July 15, 2025, the application number of 202510968820.8, and the invention name of "A method for identifying grassland grazing sheep and grassland grazing mutton and its application". TECHNICAL FIELD

[0002] The present application belongs to the technical field of food authenticity detection, and specifically relates to a method for identifying grassland grazing sheep and grassland grazing mutton and its application. BACKGROUND

[0003] In the development process of modern animal husbandry, consumers' attention to the quality and origin of mutton is increasing. Sheep meat, as an important meat product, is mainly produced in two modes: indoor feeding and grazing. Compared with indoor feeding mutton, grassland grazing mutton is favored in the market due to its unique flavor, high protein content, low fat content, and rich vitamins and minerals. It often fetches a higher price and meets the current consumer trend of pursuing healthy and natural food. However, due to the similarity in appearance between grassland grazing mutton and indoor feeding mutton, it is difficult to accurately distinguish between the two by traditional sensory identification methods such as observing meat color, texture, and touching meat quality. To effectively solve this problem, a series of methods for identifying grassland grazing mutton and indoor feeding mutton have been proposed. In terms of traditional identification methods, domestic and foreign researches have focused on the analysis of conventional physicochemical indicators of meat products. Some studies have measured and compared the color, pH value, shear force, and cooking loss of grazing and indoor feeding mutton. It was found that the color of grassland grazing mutton is usually brighter, and the pH value is relatively stable, which may be related to the large amount of exercise of grazing sheep and the consumption of natural pasture. However, these traditional methods have obvious limitations and are easily affected by various factors such as breed, age, and slight differences in feeding environment, which can cause fluctuations in physicochemical indicators and affect the accuracy and reliability of identification. At the same time, instrumental analysis techniques have also been widely used in the field of mutton identification. For example, by analyzing the spectral characteristics of mutton in the near-infrared region, the chemical composition and structure information of the meat can be obtained. However, this technique has high requirements for sample pretreatment, and the spectral data analysis process is complex and easily affected by environmental factors such as temperature and humidity, which can cause deviations in spectral data and affect the identification results. In summary, the existing methods for identifying grassland grazing mutton have certain shortcomings in accuracy, reliability, convenience, and cost-effectiveness, and cannot meet the urgent need for rapid and accurate identification of grassland grazing mutton. Therefore, developing a more accurate, reliable, efficient, and cost-effective method for identifying grassland grazing mutton is an urgent problem in the field. SUMMARY

[0004] In order to solve the problems existing in the prior art, the first objective of the present invention is to provide a grassland-grazing sheep and a method for identifying grassland-grazing mutton, with an identification accuracy of 100%.

[0005] The second objective of this invention is to provide the application of the above-mentioned method in the quality monitoring of mutton.

[0006] To achieve the above-mentioned objectives, the present invention provides the following technical solution: This invention provides a method for identifying pasture-grazing sheep, comprising the following steps: Detection of δ in sheep neck wool 13 C、δ 15 N stable isotope value; if δ is also satisfied 13 C≤-19.0‰, δ 15 If N ≥ 4.0‰, the sheep are identified as grazing sheep on grassland.

[0007] Preferably, the detection of δ in sheep neck wool 13 C、δ 15 The method for determining the stable isotope value of nitrogen includes: shearing and grinding sheep neck wool, extracting it using Soxhlet extraction with petroleum ether as solvent, defatting and drying the sample to be tested; and then measuring the sample using a stable isotope mass spectrometer.

[0008] This invention provides a method for identifying the meat of mutton from grassland-grazed sheep, comprising the following steps: Detection of δ in mutton 13 C、δ 15 N stable isotope value, content of fatty acids, flavor amino acids, and total amino acids; if δ is also satisfied 13 C≤-19.0‰, δ 15 N≥4.0‰, fatty acids ω-6:ω-3≤2.5, flavor amino acids: total amino acids≥50%, judged as grassland-grazing mutton.

[0009] Preferably, the detection of δ in mutton 13 C、δ 15 The method for determining the stable isotope value of nitrogen includes: removing fat from mutton, mincing, freeze-drying, grinding, using Soxhlet extraction with petroleum ether as solvent, defatting, and drying to obtain the sample to be tested; the sample to be tested is then determined using a stable isotope mass spectrometer.

[0010] Preferably, the amount of ω-6 in the fatty acids is the total amount of linoleic acid, γ-linolenic acid, eicosatrienoic acid, and arachidonic acid.

[0011] Preferably, the amount of ω-3 in the fatty acids is the total amount of α-linolenic acid, DHA, and EPA.

[0012] Preferably, the amount of the flavor amino acids is the total amount of glutamic acid, aspartic acid, phenylalanine, alanine, glycine, and tyrosine.

[0013] Preferably, the total amount of amino acids is the total amount of aspartic acid, threonine, serine, glutamic acid, proline, glycine, alanine, valine, methionine, isoleucine, leucine, tyrosine, phenylalanine, histidine, lysine, and arginine.

[0014] Preferably, the grassland-grazed mutton includes grassland-grazed sheep meat or grassland-grazed goat meat.

[0015] This invention also provides the application of the above-mentioned identification method in the quality monitoring of mutton.

[0016] Compared with the prior art, the beneficial effects of the technical solution of the present invention are as follows: This invention identifies the neck hair of live sheep from the perspective of stable carbon and nitrogen isotopes. It can be used to assess whether the subject is a pasture-grazing sheep, and the identification accuracy for pasture-grazing sheep reaches 99.2%.

[0017] This invention establishes a precise identification technology for grassland-grazing mutton from multiple perspectives, including carbon and nitrogen stable isotopes, the ω-6 / ω-3 fatty acid ratio, and the proportion of flavor-enhancing amino acids. It highlights the essential characteristics of grassland-grazing mutton and can eliminate the influence of factors such as breed, region, age, and human interference on grassland-grazing mutton, achieving an identification accuracy of 100%. Attached Figure Description

[0018] Figure 1 The correlation between stable C and N isotopes in mutton and stable isotopes in sheep neck hair; Figure 2 δ of neck wool from pasture-grazing sheep versus neck wool from stall-fed sheep 13 C value; Figure 3 δ of neck wool from pasture-grazing sheep versus neck wool from stall-fed sheep 15 N value; Figure 4 δ0.05 of grassland-grazed mutton versus stall-fed mutton 13 C value; Figure 5 δ0.05 of grassland-grazed mutton versus stall-fed mutton 15 N value; Figure 6 : Distribution of fatty acid ω-6 / ω-3 in grassland-grazed mutton and stall-fed mutton; Figure 7 Distribution of flavor amino acids / total amino acids in grazing-fed mutton and stall-fed mutton. Detailed Implementation

[0019] This invention provides a method for identifying pasture-grazing sheep, comprising the following steps: detecting the δ-value of sheep neck hair. 13 C、δ 15 N stable isotope value; if δ is also satisfied 13 C≤-19.0‰, δ 15 If N ≥ 4.0‰, the sheep are identified as pasture-grazing sheep. This invention utilizes the δ-value of sheep neck wool... 13 C、δ 15 The identification method of N can be used to identify sheep that are pasture-grazing before they are put on the market or before they are slaughtered, so as to identify in advance whether the sheep to be slaughtered are pasture-grazing sheep.

[0020] In this invention, the δ-value of sheep neck hair is detected. 13 C、δ 15 The method for determining stable isotope values ​​of nitrogen includes: shearing and grinding sheep neck wool, using Soxhlet extraction with petroleum ether as solvent, defatting, and air-drying to obtain the sample to be tested; the sample to be tested is then determined using a stable isotope mass spectrometer. Preferably, the sheep neck wool is sheared to 0.5 cm, ground to 100 mesh, wrapped in filter paper, and subjected to Soxhlet extraction with petroleum ether as solvent. After defatting at 60°C for 6 h, the sample is naturally air-dried to obtain the sample to be tested for stable isotopes of the hair.

[0021] This invention provides a method for identifying the meat of grazing sheep, comprising the following steps: detecting δ in the mutton. 13 C、δ 15 N stable isotope value, content of fatty acids, flavor amino acids, and total amino acids; if δ is also satisfied 13 C≤-19.0‰, δ 15 N≥4.0‰, fatty acids ω-6:ω-3≤2.5, flavor amino acids: total amino acids≥50%, judged as grassland-grazing mutton.

[0022] In this invention, δ-carboxylic acid is detected in mutton. 13 C、δ 15The method for determining stable isotope values ​​of nitrogen includes: removing fat from mutton, mincing, freeze-drying, grinding, using Soxhlet extraction with petroleum ether as solvent, defatting, and air-drying to obtain the sample to be tested; the sample to be tested is then analyzed using a stable isotope mass spectrometer. The mutton can be selected as foreleg meat. As an optional implementation method, the mutton collected on-site is sealed in self-sealing bags and stored at -20℃. Before testing, the frozen mutton is thawed at 4℃, and a partially thawed sample is taken for testing. As another optional implementation method, the present invention uses on-site samples for direct on-site testing. After removing the fat from the mutton to be tested, it is cut into small cubes, minced, and then vacuum freeze-dried. The vacuum freeze-drying conditions can be selected as -70℃ for 48 hours. The freeze-dried mutton is then pulverized and ground to 100 mesh. The sample is wrapped in filter paper and subjected to Soxhlet extraction with petroleum ether as solvent, defatted at 60℃ for 6 hours, and then naturally air-dried to obtain the sample to be tested with stable isotopes.

[0023] In this invention, the instrumentation conditions for determination using a stable isotope mass spectrometer are as follows: helium purge flow rate of 200 mL / min, combustion furnace temperature of 960℃, reduction furnace temperature of 650℃, and carrier gas He flow rate of 90~100 mL / min. The He dilution pressure is 0.6 bar, the CO2 gas pressure is 0.6 bar, and the N2 gas pressure is 1.0 bar. A USGS24 (δ) is used. 13 CO2 cylinders are calibrated using CPDB = -16.00‰, and IAEAN (δ) is used. 15 CO2 cylinders are calibrated using a calibrated cylinder gauge (Nair=0.4‰).

[0024] In this invention, the fatty acid content is preferably determined by gas chromatography, and the determination method is preferably based on GB5009.168-2016 "National Food Safety Standard - Determination of Fatty Acids in Food". The method of this invention can detect the content of 37 fatty acids. The amount of ω-6 fatty acids is the total amount of linoleic acid, γ-linolenic acid, eicosatrienoic acid, and arachidonic acid; the amount of ω-3 fatty acids is the total amount of α-linolenic acid, DHA (docosahexaenoic acid), and EPA (eicosapentaenoic acid).

[0025] In this invention, the amino acid content is preferably determined using an amino acid analyzer, and the determination method is preferably based on GB 5009.124-2016 "National Food Safety Standard - Determination of Amino Acids in Food". The method of this invention can detect the content of 16 hydrolyzed amino acids. The amount of flavor amino acids (FAA) is the total amount of glutamic acid, aspartic acid, phenylalanine, alanine, glycine, and tyrosine; the amount of total amino acids (TAA) is the total amount of aspartic acid, threonine, serine, glutamic acid, proline, glycine, alanine, valine, methionine, isoleucine, leucine, tyrosine, phenylalanine, histidine, lysine, and arginine.

[0026] In this invention, grassland-grazed mutton includes grassland-grazed sheep meat or grassland-grazed goat meat.

[0027] This invention also provides the application of the above-mentioned identification method in the quality monitoring of mutton.

[0028] The technical solutions of this invention will be clearly and completely described below with reference to the embodiments thereof. Obviously, the described embodiments are only a part of the embodiments of this invention, and not all of them. All other embodiments obtained by those skilled in the art based on the embodiments of this invention without creative effort are within the scope of protection of this invention.

[0029] Unless otherwise specified, the following embodiments are all conventional methods.

[0030] Unless otherwise specified, all materials and reagents used in the following examples are commercially available.

[0031] Example 1 A method for identifying the meat of grazing sheep: detecting δ-rays in the mutton. 13 C、δ 15 N stable isotope value, content of fatty acids, flavor amino acids, and total amino acids; if δ is also satisfied 13 C≤-19.0‰, δ 15 N≥4.0‰, fatty acids ω-6:ω-3≤2.5, flavor amino acids: total amino acids≥50%, judged as grassland-grazing mutton.

[0032] The foreleg of mutton was used as the sample for the determination of stable isotopes, fatty acids and amino acids. 2.0 kg of foreleg mutton was collected and sealed in a self-sealing bag and stored at -20℃. When the samples were ready for testing, the frozen mutton was placed at room temperature and taken out when it was partially thawed.

[0033] Detection of δ in mutton 13C、δ 15 Nitrogen stable isotope value: When testing mutton, remove the fat portion and cut the mutton into 1 cm pieces. 3 Small pieces of mutton were minced using a meat grinder and transferred to a petri dish. The minced sample was then freeze-dried at -70℃ for 48 h in a vacuum freeze dryer. After freeze-drying, the sample was pulverized and ground to 100 mesh using a ball mill. The sample was then wrapped in filter paper and extracted using the Soxhlet method with petroleum ether as the solvent. After defatting at 60℃ for 6 h, the sample was air-dried to obtain the stable isotope sample. The stable isotope mass spectrometer was used for determination. The instrument conditions were as follows: helium purge flow rate of 200 mL / min, combustion furnace temperature of 960℃, reduction furnace temperature of 650℃, and carrier gas He flow rate of 90~100 mL / min. The He dilution pressure was 0.6 bar, the CO2 reference gas pressure was 0.6 bar, and the N2 reference gas pressure was 1.0 bar. The sample was analyzed using USGS24 (δ¹⁴). 13 CO2 cylinders are calibrated using CPDB = -16.00‰, and IAEAN (δ) is used. 15 CO2 cylinders are calibrated using a calibrated cylinder apparatus (Nair=0.4‰).

[0034] To test fatty acids in mutton: cut mutton into 1 cm pieces. 3Small pieces of mutton were minced using a meat grinder and transferred to a petri dish. The minced sample was then freeze-dried at -70℃ for 48 hours in a vacuum freeze dryer. After freeze-drying, the sample was pulverized and ground to 100 mesh using a ball mill. The determination method was based on GB 5009.168-2016 "National Food Safety Standard - Determination of Fatty Acids in Food". The specific procedure was as follows: 0.5 g of sample was weighed and transferred to a 250 mL flat-bottomed flask. 2.0 mL of undecylate triglyceride internal standard solution was accurately added, along with approximately 200 mg of pyrogallol, a few boiling chips, 2 mL of 95% ethanol, and 4 mL of water. The mixture was thoroughly mixed, and 10 mL of 8.3 mol / L hydrochloric acid solution was added and mixed well. The flask was then placed in an 80℃ water bath for hydrolysis for 40 min. After hydrolysis, the flask was removed and cooled to room temperature. After hydrolysis, add 10 mL of 95% ethanol, mix well, and transfer the hydrolysate from the flask to a separatory funnel. Rinse the flask and stopper with 50 mL of diethyl ether and petroleum ether (volume ratio 1:1). Add the rinsing solution to the separatory funnel, cap, shake for 5 min, and let stand for 10 min. Collect the ether extract in a 250 mL flask. Repeat the above steps three times. Combine the ether extracts and concentrate to dryness using a rotary evaporator to obtain the fat extract. Add 8 mL of 2% sodium hydroxide methanol solution to the fat extract, connect a reflux condenser, and reflux at 80°C in a water bath until the oil droplets disappear. Add 7 mL of 15% boron trifluoride methanol solution from the top of the reflux condenser and continue reflux at 80°C for 2 min. Rinse the reflux condenser with a small amount of water. Stop heating, remove the flask from the water bath, and quickly cool to room temperature. Accurately add 20 mL of n-heptane, shake for 2 min, then add saturated sodium chloride aqueous solution and let stand to separate the layers. Pipette 5 mL of the upper n-heptane extract into a 25 mL test tube, add 5 g of anhydrous sodium sulfate, shake for 1 min, let stand for 5 min, and then transfer the upper layer solution to a sample vial for analysis. Gas chromatography conditions: column (polydicyanopropylsiloxane strongly polar stationary phase, column length 100 m, inner diameter 0.25 mm, film thickness 0.2 μm), injector temperature 270 ℃, detector temperature 280 ℃, carrier gas N2, split ratio 100:1, injection volume 100:1.

[0035] Testing for amino acids in mutton: Cut the mutton into 1 cm pieces. 3Small pieces of mutton were minced using a meat grinder and transferred to a petri dish. The minced sample was then freeze-dried at -70℃ for 48 hours in a vacuum freeze dryer. After freeze-drying, the sample was pulverized and ground to 100 mesh using a ball mill. The determination method was based on GB 5009.124-2016 "National Food Safety Standard - Determination of Amino Acids in Food". The specific procedure was as follows: 0.5 g of the sample was accurately weighed and placed in a hydrolysis tube. 15 mL of 6 mol / L hydrochloric acid solution was added, along with 3-4 drops of phenol. The hydrolysis tube was frozen with liquid nitrogen for 3 minutes, then evacuated and purged with nitrogen. This process was repeated 3 times. The tube was sealed under nitrogen purging and placed in a 110℃ electric heating drying oven for 22 hours for hydrolysis. After removal, the tube was cooled to room temperature. The hydrolysate was filtered into a 50 mL volumetric flask, diluted to the mark with water, and shaken well. This solution was used for the determination of amino acids. Instrument testing conditions: The chromatographic column is a sulfonic acid type cation exchange resin, and the detection wavelengths are 570 nm and 440 nm.

[0036] Example 2 A method for identifying pasture-grazing sheep: detecting the δ value of sheep neck wool. 13 C、δ 15 N stable isotope value; if δ is also satisfied 13 C≤-19.0‰, δ 15 If N ≥ 4.0‰, the sheep are identified as grazing sheep on grassland.

[0037] Sheep neck hair was collected as a sample for stable isotope determination. The δ¹⁸O₂ concentration in the sheep neck hair was detected. 13 C、δ 15 Stable isotope values ​​of nitrogen (δ¹⁸O) were obtained by collecting sheep neck hair, cutting it to 0.5 cm, grinding it to 100 mesh using a ball mill, wrapping the sample in filter paper, and using Soxhlet extraction with petroleum ether as solvent. After defatting at 60°C for 6 h, the sample was air-dried to obtain the hair stable isotope sample. The stable isotope values ​​were determined using a stable isotope mass spectrometer, with the instrument conditions the same as described in Example 1 for "Detection of δ¹⁸O in mutton". 13 C、δ 15 The "stable isotope values ​​of N" are the same.

[0038] Experimental Example 1 With the δ of sheep neck wool 13 C and δ 15 N is a technical indicator, and its formation and establishment are based on the following: Samples of neck hair and corresponding mutton from grazing sheep on grasslands are collected, and the δ0 values ​​of the mutton and neck hair are measured. 13 C and δ 15 The N-value was used to establish a linear model of sheep neck wool and mutton using correlation analysis, revealing the δ-value in mutton. 13 C、δ 15 N and sheep neck wool δ 13 C、δ 15The degree of correlation of N. Lamb δ 13 C and δ 15 The method for determining N is the same as in Example 1, and the δ of sheep neck wool is... 13 C and δ 15 The method for determining N is the same as in Example 2.

[0039] The results are as follows Figure 1 As shown, the results indicate that δ-carboxylic acid in mutton... 13 C and δ in sheep neck wool 13 C showed a highly significant positive correlation with 0.917, and the δ in mutton... 15 N and δ of sheep neck hair 15 N showed a highly significant positive correlation, with a correlation coefficient of 0.9414. Therefore, sheep neck wool was included in the technical indicator system for identifying mutton from grassland-grazing sheep.

[0040] Experimental Example 2 1. 180 samples of sheep neck wool were collected, including 80 samples from pasture-grazing sheep and 100 samples from stall-fed sheep. The pasture-grazing sheep samples covered the four major pastoral areas of China (Inner Mongolia, Xinjiang, Tibet, Qinghai, Chongqing, and Yunnan) and the grassland and hillside grazing areas in southern China. The stall-fed sheep samples covered typical agricultural areas in China (Hebei, Shanxi, Inner Mongolia, and Shandong) and stall-fed areas. The carbon stable isotope (δ¹⁸O) analysis was performed using the method described in Example 2. 13 C) and nitrogen-stable isotopes (δ) 15 N) was tested, and the results were as follows: Figures 2-3 As shown.

[0041] For carbon stable isotopes (δ 13 C) measurements showed that the neck wool δ of grassland-grazing sheep... 13 The maximum value of C was -19.05‰, the minimum value was -24.0‰, the average value was -21.36‰, and the median value was -21.32‰; while the neck wool of stall-fed sheep had a δ 13 The maximum value of C is -11.83‰, the minimum value is -18.44‰, the average value is -15.45‰, and the median value is -15.71‰.

[0042] For nitrogen stable isotopes (δ 15 Measurements of N) showed that the neck wool δ of grazing sheep was... 15 The maximum value of N was 12.0‰, the minimum value was 4.0‰, the average value was 5.0‰, and the median value was 6.1‰; while the δ of neck wool in stall-fed sheep was... 15 The maximum value of N is 9.8‰, the minimum value is 1.8‰, the average value is 4.5‰, and the median value is 6.5‰.

[0043] 2. 180 mutton samples were collected, including 100 samples of grassland-grazed mutton and 80 samples of stall-fed mutton. The grassland-grazed mutton samples covered the four major pastoral areas of China, namely Inner Mongolia, Xinjiang, Tibet, Qinghai, Chongqing, and Yunnan, as well as grassland and hillside grazing areas in southern China. The stall-fed mutton samples covered typical agricultural areas in China, such as Hebei, Shanxi, Inner Mongolia, and Shandong, where stall-fed mutton was raised. The carbon stable isotope (δ¹²) analysis was performed using the method described in Example 1. 13 C) and nitrogen-stable isotopes (δ) 15 N) was tested, and the results were as follows: Figures 4-5 As shown.

[0044] For carbon stable isotopes (δ 13 C) measurements showed that the δ-carbohydrate content of grazing sheep was higher than that of other species. 13 The maximum value of C was -19.27‰, the minimum value was -24.15‰, the average value was -21.76‰, and the median value was -21.88‰; while the δ of intensively raised mutton was... 13 The maximum value of C is -13.5‰, the minimum value is -18.36‰, the average value is -15.79‰, and the median value is -15.92‰.

[0045] For nitrogen stable isotopes (δ 15 Measurements of N) showed that grassland-grazing mutton δ 15 The maximum value of N was 9.8‰, the minimum value was 4.0‰, the average value was 7.0‰, and the median value was 8.0‰; while the δ of intensively raised mutton was... 15 The maximum value of N is 8.8‰, the minimum value is 1.6‰, the average value is 5.3‰, and the median value is 7.1‰.

[0046] 3. 180 mutton samples were collected, including 100 samples of grassland-grazed mutton and 80 samples of stall-fed mutton. The grassland-grazed mutton samples covered the four major pastoral areas of China (Inner Mongolia, Xinjiang, Tibet, Qinghai, Chongqing, and Yunnan) and grassland-slope grazing areas in southern China. The stall-fed mutton samples covered typical agricultural areas of China (Hebei, Shanxi, Inner Mongolia, and Shandong) and stall-fed areas. The fatty acid content was determined and analyzed using the method described in Example 1. The results are as follows: Figure 6 As shown.

[0047] The results showed that there was a significant difference in the ω-6 / ω-3 ratio between grassland-grazed and stall-fed mutton. The maximum ω-6 / ω-3 ratio for grassland-grazed mutton was 3.01, the minimum was 0.71, the average was 1.87, and the median was 1.77. In contrast, the maximum ω-6 / ω-3 ratio for stall-fed mutton was 10.9, the minimum was 3.1, the average was 8.17, and the median was 6.66.

[0048] 4. A total of 180 mutton samples were collected, including 100 samples of grassland-grazed mutton and 80 samples of stall-fed mutton. The grassland-grazed mutton samples covered the four major pastoral areas of China (Inner Mongolia, Xinjiang, Tibet, Qinghai, Chongqing, and Yunnan) and grassland-slope grazing areas in southern China. The stall-fed mutton samples covered typical agricultural areas of China (Hebei, Shanxi, Inner Mongolia, and Shandong) and stall-fed mutton farming areas. The amino acid content was determined and analyzed using the method described in Example 1. The results are as follows: Figure 7 As shown.

[0049] The results showed that the maximum FAA / TAA ratio for grassland-grazed mutton was 57.52%, the minimum was 47.19%, the average was 52.00%, and the median was 52.68%, while the maximum FAA / TAA ratio for stall-fed mutton was 56.27%, the minimum was 45.38%, the median was 49.93%, and the average was 49.97%.

[0050] 5. SPSS software was used to perform classification and discriminant analysis on the samples, and the results are shown in Table 1.

[0051] Table 1. Discriminant analysis results of grassland-grazed mutton versus stall-fed mutton under different indicators

[0052] The results show that when using a single indicator for discrimination, δ 13 C,δ 13 C and δ 15 The original validation rate and cross-validation rate of the N, ω-6 / ω-3 fatty acid ratio reached over 90%, and the original validation rate and cross-validation rate of the FAA / TAA amino acid composition reached over 70%; when using combined indicators, i.e., δ 13 C and δ 15 When N, ω-6 / ω-3, and FAA / TAA are used for simultaneous discrimination, the original validation rate and cross-validation rate are significantly improved, both reaching 100%.

[0053] Experimental Example 3 120 samples of sheep neck wool were randomly collected, including 60 samples from pasture-grazing sheep and 60 samples from stall-fed sheep. The samples were uniformly numbered, their information summarized, and they were categorized according to their feeding type based on the actual sampling conditions. The samples were then sent to a third-party laboratory for C and N isotope testing and analysis, using the delta-grain density (δ¹⁸O) of the sheep neck wool as the analytical parameter. 13 C and δ 15 The experimental test results of N were used to verify whether the sample category was consistent with the actual situation of sample collection. The technical indicators of Example 2 were used as the identification criteria for grassland grazing sheep and stall-fed sheep to realize the verification and application of the grassland grazing sheep identification method. The results are shown in Table 2.

[0054] Table 2. Results of using C and N isotopes of sheep neck wool as a method for identifying pasture-grazing sheep.

[0055] One hundred mutton samples were collected nationwide, including 50 samples from grassland-fed mutton and 50 samples from stall-fed mutton. These samples were uniformly numbered and their information compiled before being sent to the laboratory for δ¹⁸O₂ analysis. 13 C and δ 15 The testing methods for N, fatty acid ratio, and amino acid percentage were the same as in Example 1. The technical indicators of Example 1 were used as the identification standards for grassland-grazed sheep and stall-fed mutton to verify and apply the identification method for grassland-grazed mutton. The results are shown in Table 3.

[0056] Table 3. Results of the application of identification methods for grazing mutton.

[0057] The results showed that, with a single δ 13 Using C as the identification standard yields an accuracy of 94%; using the ω-6 / ω-3 ratio as the identification standard yields an accuracy of 93%; and using the FAA / TAA ratio as the identification standard yields an accuracy of 70%. Using δ... 13 C and δ 15 Using N, ω-6 / ω-3, and the FAA / TAA combination as the identification criteria, the results had an accuracy of 100%.

[0058] The above description is only a preferred embodiment of the present invention. It should be noted that for those skilled in the art, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. A method for identifying sheep grazed on grasslands, characterized in that, Includes the following steps: Detection of δ in sheep neck wool 13 C、δ 15 N stable isotope value; if δ is also satisfied 13 C≤-19.0‰, δ 15 If N ≥ 4.0‰, the sheep are identified as grazing sheep on grassland.

2. The identification method according to claim 1, characterized in that, Detection of δ in sheep neck wool 13 C、δ 15 The method for determining the stable isotope value of nitrogen includes: shearing and grinding sheep neck wool, extracting it using Soxhlet extraction with petroleum ether as solvent, defatting and drying the sample to be tested; and then measuring the sample using a stable isotope mass spectrometer.

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