Method for detecting flavor substances in mutton sheep muscles based on mass spectrum

Abstract

The invention provides a method for detecting flavor substances in mutton sheep muscle based on mass spectrum, which comprises the following steps: selecting healthy mutton sheep, treating longissimus dorsi muscle, and quickly freezing and storing the longissimus dorsi muscle. Grinding the sample by liquid nitrogen, ultrasonically extracting flavor substances by using a methanol-water mixed solution with the volume ratio of 3: 2, centrifuging, and combining supernate; then purifying through a C18 solid-phase extraction column, and concentrating and redissolving to obtain a to-be-detected sample solution. Gas chromatography-mass spectrometry (GC-MS) and liquid chromatography-mass spectrometry (LC-MS) are adopted for detection, the GC-MS is used for qualitative determination of volatile flavor substances and quantification of an external standard method, and the LC-MS is used for qualitative determination of non-volatile flavor substances and quantification of an internal standard method. The method can effectively destroy cell structures and fully extract flavor substances, impurities can be removed in the purification process, the composition and content of the flavor substances are comprehensively and accurately analyzed by combining two mass spectrometry technologies, and powerful technical support is provided for mutton quality evaluation and flavor regulation and control.

Description

Technical Field

[0001] The present invention relates to the technical field of food detection, and more particularly, to a method for detecting flavor substances in mutton sheep muscle based on mass spectrometry. Background Art

[0002] Mutton is one of the important meat sources in people's daily life. The flavor substances in mutton sheep muscle are the key factors affecting the quality of mutton and the acceptance of consumers. There are many kinds of flavor substances, including volatile compounds, non-volatile compounds, etc. The composition and content of these substances determine the unique flavor characteristics of mutton.

[0003] At present, there are various detection methods for flavor substances in mutton sheep muscle, such as gas chromatography-mass spectrometry (GC-MS), liquid chromatography-mass spectrometry (LC-MS), etc. However, the existing detection methods have some deficiencies. On the one hand, it is difficult to fully extract all flavor substances during the sample pretreatment process, resulting in the possible failure to detect some substances with low content but important contributions to the flavor; on the other hand, the sensitivity and accuracy of the detection process need to be improved, and it is impossible to accurately perform qualitative and quantitative analysis on flavor substances. Therefore, we propose a method for detecting flavor substances in mutton sheep muscle based on mass spectrometry. Summary of the Invention

[0004] The purpose of the present invention is to address the problems raised in the existing background art. To achieve the above-mentioned invention purpose, the present invention provides the following technical solutions: A method for detecting flavor substances in mutton sheep muscle based on mass spectrometry, comprising the following steps: Step 1, sample collection and pretreatment: Select healthy mutton sheep, quickly take the longissimus dorsi muscle sample after slaughtering, rinse the surface with physiological saline to remove blood stains and impurities, then dry the surface moisture with filter paper, cut the processed muscle sample into small pieces, quickly freeze it in liquid nitrogen, and then transfer it to a -80°C refrigerator for storage and standby; Step 2, flavor substance extraction: Take out the muscle sample from the -80°C refrigerator, weigh the sample and put it into a mortar, add liquid nitrogen, and quickly grind it into a powder. Transfer the ground powder to a centrifuge tube, add an extraction solvent, vortex for 5 minutes to fully mix the sample with the extraction solvent. Place the centrifuge tube in an ultrasonic extractor and ultrasonically extract at 40°C for 30 minutes. After the ultrasonic extraction is completed, centrifuge the centrifuge tube at 12000 r / min for 15 minutes, take the supernatant and transfer it to a new centrifuge tube, and repeat the extraction of the residue with the same volume of extraction solvent once. Combine the supernatants extracted twice; Step 3, purification treatment: Purify the combined supernatant through a solid-phase extraction column; Step 4, Concentration and Reconstitution: Concentrate the eluate to near dryness at 40°C using a rotary evaporator, then reconstitute it with 1 mL of methanol and filter through a 0.22 μm organic filter membrane to obtain the sample solution to be tested; Step 5, Mass Spectrometry Detection: Detect the sample solution to be tested using gas chromatography-mass spectrometry (GC-MS) and liquid chromatography-mass spectrometry (LC-MS).

[0005] As a preferred technical solution of the present invention, the extraction solvent is a methanol-water mixed solution with a volume ratio of 3:2.

[0006] As a preferred technical solution of the present invention, the solid-phase extraction column is a C18 column. The purification process is to first activate it with 5 mL of methanol, then rinse and balance it with 5 mL of water. Pass the supernatant through the activated C18 column at a flow rate of 1 mL / min, then rinse the column with 5 mL of water to remove impurities, and finally elute the flavor substances adsorbed on the column with 5 mL of methanol and collect the eluate.

[0007] As a preferred technical solution of the present invention, the gas chromatography conditions for the gas chromatography-mass spectrometry (GC-MS) detection are as follows: the chromatographic column is an HP-5MS capillary column (30 m × 0.25 mm × 0.25 μm); the inlet temperature is 250°C; the carrier gas is helium with a flow rate of 1 mL / min; the split ratio is 10:1; the column temperature program: the initial temperature is 40°C, held for 3 minutes, heated at a rate of 5°C / min to 200°C, held for 5 minutes, and then heated at a rate of 10°C / min to 280°C, held for 5 minutes.

[0008] As a preferred technical solution of the present invention, the mass spectrometry conditions for the gas chromatography-mass spectrometry (GC-MS) detection are as follows: the ion source is an EI source, the ion source temperature is 230°C; the interface temperature is 280°C; the scanning range is m / z 35 - 500.

[0009] As a preferred technical solution of the present invention, in the gas chromatography-mass spectrometry (GC-MS) detection, inject 1 μL of the sample solution to be tested into the gas chromatograph, and perform qualitative analysis of the volatile flavor substances by comparing with the NIST mass spectrometry database, and perform quantitative analysis of the volatile flavor substances using the external standard method.

[0010] As a preferred technical solution of the present invention, the liquid chromatography conditions for the liquid chromatography-mass spectrometry (LC-MS) detection are as follows: the chromatographic column is a C18 reverse-phase chromatographic column (250 mm × 4.6 mm, 5 μm); mobile phase A is an aqueous solution of 0.1% formic acid, and mobile phase B is acetonitrile; gradient elution program: 0 - 5 minutes, 5% mobile phase B; 5 - 20 minutes, 5% - 95% mobile phase B; 20 - 25 minutes, 95% mobile phase B; 25 - 30 minutes, 95% - 5% mobile phase B; the flow rate is 1 mL / min; the column temperature is 30°C.

[0011] As a preferred technical solution of the present invention, the mass spectrometry conditions for the liquid chromatography-mass spectrometry (LC-MS) detection are as follows: the ion source is an ESI source, in positive ion mode; the spray voltage is 3.5 kV; the capillary temperature is 320°C; the scanning range is m / z 100 - 1000.

[0012] As a preferred technical solution of the present invention, in the liquid chromatography-mass spectrometry (LC-MS) detection, 10 μL of the sample solution to be measured is injected into the liquid chromatograph, and the non-volatile flavor substances are qualitatively analyzed by comparing the retention time and mass spectrometry diagram with the standard product, and the non-volatile flavor substances are quantitatively analyzed by the internal standard method.

[0013] As a preferred technical solution of the present invention, in the sample collection and pretreatment steps, the frozen muscle samples are stored in a -80°C refrigerator for a long time.

[0014] Compared with the prior art, the beneficial effects of the present invention are as follows: in the solution of the present invention: the present invention adopts the method of grinding with liquid nitrogen to pretreat the muscle samples, which can effectively destroy the cell structure, fully release the flavor substances, and improve the extraction efficiency of the flavor substances.

[0015] Select a methanol-water mixed solution with a volume ratio of 3:2 as the extraction solvent, which has good solubility for both volatile and non-volatile flavor substances, and can comprehensively extract the flavor substances in mutton sheep muscle.

[0016] The extract is purified by a solid-phase extraction column, which can remove impurities and interfering substances in the sample and improve the sensitivity and accuracy of the detection.

[0017] Combining two detection techniques of gas chromatography-mass spectrometry (GC-MS) and liquid chromatography-mass spectrometry (LC-MS) to detect volatile and non-volatile flavor substances respectively can comprehensively and accurately analyze the composition and content of flavor substances in mutton sheep muscle. Description of the Drawings

[0018] Figure 1 It is a test data block diagram provided by the present invention; Figure 2The block diagram of the test data provided by the present invention; Figure 3 The flowchart of the method provided by the present invention. Detailed implementation manners

[0019] To make the objectives, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are some but not all of the embodiments of the present invention.

[0020] Therefore, the following detailed description of the embodiments of the present invention is not intended to limit the scope of the claimed invention, but merely represents some embodiments of the present invention. All other embodiments obtained by those of ordinary skill in the art based on the embodiments of the present invention without creative efforts fall within the scope of protection of the present invention. It should be noted that, without conflict, the embodiments in the present invention and the features and technical solutions in the embodiments may be combined with each other. It should be noted that similar reference numerals and letters denote similar items in the following drawings. Therefore, once an item is defined in one drawing, it does not need to be further defined and explained in subsequent drawings.

[0021] Embodiment 1: A method for detecting flavor substances in mutton sheep muscle based on mass spectrometry, comprising the following steps: Step 1, sample collection and pretreatment: Select healthy mutton sheep, quickly take the longissimus dorsi muscle samples after slaughtering, rinse the surface with physiological saline to remove blood stains and impurities, then dry the surface moisture with filter paper, cut the processed muscle samples into small pieces, quickly freeze them in liquid nitrogen, and then transfer them to a -80°C refrigerator for storage for later use; Step 2, flavor substance extraction: Take out the muscle samples from the -80°C refrigerator, weigh the samples and put them into a mortar, add liquid nitrogen, and quickly grind them into powder. Transfer the ground powder to a centrifuge tube, add an extraction solvent, vortex for 5 minutes to fully mix the samples with the extraction solvent, place the centrifuge tube in an ultrasonic extractor, ultrasonically extract at 40°C for 30 minutes. After the ultrasonic extraction is completed, centrifuge the centrifuge tube at 12000 r / min for 15 minutes, take the supernatant and transfer it to a new centrifuge tube, and repeat the extraction of the residue once with the same volume of extraction solvent, and combine the supernatants extracted twice; Step 3, purification treatment: Purify the combined supernatant through a solid-phase extraction column; Step 4, concentration and reconstitution: Concentrate the eluate to near dryness at 40°C with a rotary evaporator, then reconstitute it with 1 mL of methanol, and filter it through a 0.22 μm organic filter membrane to obtain a sample solution to be tested; Step 5, mass spectrometry detection: Use gas chromatography-mass spectrometry (GC-MS) and liquid chromatography-mass spectrometry (LC-MS) to detect the sample solution to be tested.

[0022] The extraction solvent is a methanol-water mixed solution with a volume ratio of 3:2.

[0023] The solid-phase extraction column is a C18 column. The purification process is as follows: first, activate it with 5 mL of methanol, then rinse and balance it with 5 mL of water. Pass the supernatant through the activated C18 column at a flow rate of 1 mL / min. Then, rinse the column with 5 mL of water to remove impurities. Finally, elute the flavor substances adsorbed on the column with 5 mL of methanol and collect the eluate.

[0024] The gas chromatographic conditions for gas chromatography-mass spectrometry (GC-MS) detection are as follows: the chromatographic column is an HP-5MS capillary column (30 m × 0.25 mm × 0.25 μm); the inlet temperature is 250 °C; the carrier gas is helium with a flow rate of 1 mL / min; the split ratio is 10:1; the column temperature program: the initial temperature is 40 °C, hold for 3 minutes, increase the temperature to 200 °C at a rate of 5 °C / min, hold for 5 minutes, and then increase the temperature to 280 °C at a rate of 10 °C / min, hold for 5 minutes.

[0025] The mass spectrometric conditions for gas chromatography-mass spectrometry (GC-MS) detection are as follows: the ion source is an EI source, and the ion source temperature is 230 °C; the interface temperature is 280 °C; the scanning range is m / z 35 - 500.

[0026] In gas chromatography-mass spectrometry (GC-MS) detection, inject 1 μL of the sample solution to be measured into the gas chromatograph. Qualitative analysis of the volatile flavor substances is carried out by comparing with the NIST mass spectrometry database, and quantitative analysis of the volatile flavor substances is carried out by the external standard method.

[0027] The liquid chromatographic conditions for liquid chromatography-mass spectrometry (LC-MS) detection are as follows: the chromatographic column is a C18 reversed-phase chromatographic column (250 mm × 4.6 mm, 5 μm); mobile phase A is an aqueous solution of 0.1% formic acid, and mobile phase B is acetonitrile; the gradient elution program: 0 - 5 minutes, 5% mobile phase B; 5 - 20 minutes, 5% - 95% mobile phase B; 20 - 25 minutes, 95% mobile phase B; 25 - 30 minutes, 95% - 5% mobile phase B; the flow rate is 1 mL / min; the column temperature is 30 °C.

[0028] The mass spectrometric conditions for liquid chromatography-mass spectrometry (LC-MS) detection are as follows: the ion source is an ESI source, in positive ion mode; the spray voltage is 3.5 kV; the capillary temperature is 320 °C; the scanning range is m / z 100 - 1000.

[0029] In the liquid chromatography-mass spectrometry (LC-MS) detection, 10 μL of the sample solution to be measured is injected into the liquid chromatograph. By comparing the retention time and mass spectrum with the standard, qualitative analysis of non-volatile flavor substances is carried out, and quantitative analysis of non-volatile flavor substances is carried out by the internal standard method.

[0030] In the sample collection and pretreatment steps, the frozen muscle samples are stored in a -80 °C refrigerator for a long time.

[0031] Example 2: A method for detecting flavor substances in mutton sheep muscle based on mass spectrometry, comprising the following steps: Sample collection and pretreatment Select healthy mutton sheep. After slaughter, quickly take the longissimus dorsi muscle samples, rinse the surface with physiological saline to remove blood stains and impurities, and then blot the surface moisture with filter paper.

[0032] Cut the processed muscle samples into small pieces, freeze them quickly in liquid nitrogen, and then transfer them to a -80 °C refrigerator for storage for later use.

[0033] Flavor substance extraction Take out the muscle samples from the -80 °C refrigerator, weigh the samples and put them into a mortar, add liquid nitrogen, and quickly grind them into powder.

[0034] Transfer the ground powder to a centrifuge tube, add the extraction solvent (in this invention, a methanol-water mixed solution with a volume ratio of 3:2 is used), vortex for 5 minutes to fully mix the sample with the extraction solvent.

[0035] Place the centrifuge tube in an ultrasonic extractor and ultrasonically extract at 40 °C for 30 minutes to promote the dissolution of flavor substances.

[0036] After the ultrasonic extraction is completed, centrifuge the centrifuge tube at 12000 r / min for 15 minutes, and transfer the supernatant to a new centrifuge tube.

[0037] Repeat the extraction of the residue once with the same volume of extraction solvent, and combine the supernatants extracted twice.

[0038] Purification treatment Purify the combined supernatant through a solid-phase extraction column. The solid-phase extraction column used in this invention is a C18 column. First, activate it with 5 mL of methanol, and then rinse and balance it with 5 mL of water.

[0039] Pass the supernatant through the activated C18 column at a flow rate of 1 mL / min, and then rinse the column with 5 mL of water to remove impurities.

[0040] Finally, elute the flavor substances adsorbed on the column with 5 mL of methanol, and collect the eluate.

[0041] Concentration and reconstitution The eluate was concentrated to near dryness at 40 °C using a rotary evaporator, then redissolved in 1 mL of methanol, and filtered through a 0.22 μm organic filter membrane to obtain the sample solution to be tested.

[0042] Mass spectrometry detection Gas chromatography - mass spectrometry (GC - MS) detection Gas chromatography conditions: The chromatographic column was an HP - 5MS capillary column (30 m × 0.25 mm × 0.25 μm); the inlet temperature was 250 °C; the carrier gas was helium with a flow rate of 1 mL / min; the split ratio was 10:1; the column temperature program: the initial temperature was 40 °C, held for 3 minutes, heated at a rate of 5 °C / min to 200 °C, held for 5 minutes, then heated at a rate of 10 °C / min to 280 °C, held for 5 minutes.

[0043] Mass spectrometry conditions: The ion source was an EI source, the ion source temperature was 230 °C; the interface temperature was 280 °C; the scanning range was m / z 35 - 500.

[0044] 1 μL of the sample solution to be tested was injected into the gas chromatograph for GC - MS analysis. The volatile flavor substances were qualitatively analyzed by comparing with the NIST mass spectrometry database, and the volatile flavor substances were quantitatively analyzed by the external standard method.

[0045] Liquid chromatography - mass spectrometry (LC - MS) detection Liquid chromatography conditions: The chromatographic column was a C18 reversed - phase chromatographic column (250 mm × 4.6 mm, 5 μm); mobile phase A was 0.1% formic acid aqueous solution, mobile phase B was acetonitrile; gradient elution program: 0 - 5 minutes, 5% mobile phase B; 5 - 20 minutes, 5% - 95% mobile phase B; 20 - 25 minutes, 95% mobile phase B; 25 - 30 minutes, 95% - 5% mobile phase B; the flow rate was 1 mL / min; the column temperature was 30 °C.

[0046] Mass spectrometry conditions: The ion source was an ESI source, positive ion mode; the spray voltage was 3.5 kV; the capillary temperature was 320 °C; the scanning range was m / z 100 - 1000.

[0047] 10 μL of the sample solution to be tested was injected into the liquid chromatograph for LC - MS analysis. The non - volatile flavor substances were qualitatively analyzed by comparing the retention time and mass spectrometry diagram with the standard product, and the non - volatile flavor substances were quantitatively analyzed by the internal standard method.

[0048] Experimental Example 1: Materials and Methods Experimental materials: 10 mutton sheep of the same breed, healthy and with similar body weights (about 30 - 35 kg), were selected. After slaughter, the longissimus dorsi muscle samples were quickly taken and processed and detected according to the method described in the invention.

[0049] Test method: Operate according to "A method for detecting flavor substances in mutton sheep muscle based on mass spectrometry", including sample collection and pretreatment, flavor substance extraction, purification treatment, concentration and reconstitution, and mass spectrometry detection (GC-MS and LC-MS).

[0050] Test process: Process 10 muscle samples in sequence to ensure that the operation of each step is standardized and consistent.

[0051] During the GC-MS and LC-MS detection processes, analyze strictly according to the set instrument conditions.

[0052] Test data table: Table 1: Results of GC-MS detection of volatile flavor substances (mg / kg)

[0053] Table 2: Results of LC-MS detection of non-volatile flavor substances (mg / kg)

[0054] Data analysis Volatile flavor substances Statistically analyze the data of aldehydes, ketones, alcohols and other volatile flavor substances in Table 1. Calculate the average value and standard deviation of the content of each substance. The results show that the average content of aldehydes is 1.23 ± 0.04 mg / kg, the average content of ketones is 0.90 ± 0.03 mg / kg, the average content of alcohols is 0.57 ± 0.03 mg / kg, and the average content of other types is 0.22 ± 0.01 mg / kg. The content of each substance fluctuates little among different samples, indicating that this detection method has good stability for the detection of volatile flavor substances.

[0055] Non-volatile flavor substances Statistically analyze the data of amino acids, nucleotides, organic acids and other non-volatile flavor substances in Table 2. The average content of amino acids is 15.21 ± 0.20 mg / kg, the average content of nucleotides is 8.62 ± 0.12 mg / kg, the average content of organic acids is 3.22 ± 0.04 mg / kg, and the average content of other types is 1.13 ± 0.02 mg / kg. Similarly, the content of each substance fluctuates little among different samples, indicating that this method also has good stability for the detection of non-volatile flavor substances.

[0056] Conclusion The method for detecting flavor substances in mutton sheep muscle based on mass spectrometry provided by the present invention can effectively detect volatile and non-volatile flavor substances in mutton sheep muscle.

[0057] Through the analysis of the test data of 10 samples, it can be seen that this test method has good stability and repeatability, and can provide reliable data support for the research of mutton sheep muscle flavor substances and the evaluation of mutton quality.

[0058] This method is feasible in practical applications and can be used for the monitoring and control of mutton flavor quality in the links of mutton sheep breeding, slaughtering and processing.

[0059] The above embodiments are only used to illustrate the present invention and do not limit the technical solutions described in the present invention. Although this specification has described the present invention in detail with reference to the above embodiments, the present invention is not limited to the above specific embodiments. Therefore, any modification or equivalent replacement of the present invention; and all technical solutions and their improvements that do not depart from the spirit and scope of the invention are covered by the scope of the claims of the present invention.

Claims

1. A method for detecting flavor substances in mutton sheep muscle based on mass spectrometry, characterized in that, It includes the following steps: Step 1, sample collection and pretreatment: Select healthy mutton sheep. After slaughter, quickly take the longissimus dorsi muscle samples. Rinse the surface with physiological saline to remove blood stains and impurities, then blot the surface moisture with filter paper. Cut the treated muscle samples into small pieces, quickly freeze them in liquid nitrogen, and then transfer them to a -80°C refrigerator for storage and standby; Step 2, flavor substance extraction: Take out the muscle samples from the -80°C refrigerator, weigh the samples and put them into a mortar, add liquid nitrogen, and quickly grind them into powder. Transfer the ground powder to a centrifuge tube, add the extraction solvent, vortex for 5 minutes to fully mix the sample with the extraction solvent. Place the centrifuge tube in an ultrasonic extractor and ultrasonically extract at 40°C for 30 minutes. After the ultrasonic extraction, centrifuge the centrifuge tube at 12000 r / min for 15 minutes, take the supernatant and transfer it to a new centrifuge tube. Repeat the extraction of the residue once with the same volume of extraction solvent, and combine the supernatants of the two extractions; Step 3, purification treatment: Purify the combined supernatant through a solid-phase extraction column; Step 4, concentration and reconstitution: Concentrate the eluate to nearly dry at 40°C using a rotary evaporator, then reconstitute it with 1 mL of methanol, and filter it through a 0.22 μm organic filter membrane to obtain the sample solution to be tested; Step 5, mass spectrometry detection: Detect the sample solution to be tested by gas chromatography-mass spectrometry (GC-MS) and liquid chromatography-mass spectrometry (LC-MS).

2. The method for detecting flavor substances in mutton sheep muscle based on mass spectrometry according to claim 1, wherein, The extraction solvent is a methanol-water mixed solution with a volume ratio of 3:

2.

3. The method for detecting flavor substances in mutton sheep muscle based on mass spectrometry according to claim 1, characterized in that The solid-phase extraction column is a C18 column. The purification process is to first activate it with 5 mL of methanol, then rinse and balance it with 5 mL of water. Pass the supernatant through the activated C18 column at a flow rate of 1 mL / min, then rinse the column with 5 mL of water to remove impurities, and finally elute the flavor substances adsorbed on the column with 5 mL of methanol, and collect the eluate.

4. The method for detecting flavor substances in mutton sheep muscle based on mass spectrometry according to claim 1, wherein The gas chromatography conditions for the gas chromatography-mass spectrometry (GC-MS) detection are as follows: The chromatographic column is an HP-5MS capillary column (30 m × 0.25 mm × 0.25 μm); the injection port temperature is 250°C; the carrier gas is helium with a flow rate of 1 mL / min; the split ratio is 10:1; the column temperature program: the initial temperature is 40°C, hold for 3 minutes, increase the temperature to 200°C at a rate of 5°C / min, hold for 5 minutes, then increase the temperature to 280°C at a rate of 10°C / min, and hold for 5 minutes.

5. The method for detecting flavor substances in mutton sheep muscle based on mass spectrometry according to claim 1, wherein The mass spectrometry conditions for the gas chromatography-mass spectrometry (GC-MS) detection are as follows: The ion source is an EI source, the ion source temperature is 230°C; the interface temperature is 280°C; the scanning range is m / z 35 - 500.

6. The method for detecting flavor substances in mutton sheep muscle based on mass spectrometry according to claim 1, wherein, In the gas chromatography-mass spectrometry (GC-MS) detection, inject 1 μL of the sample solution to be tested into the gas chromatograph. Qualitatively analyze the volatile flavor substances by comparing with the NIST mass spectrometry database, and quantitatively analyze the volatile flavor substances by the external standard method.

7. The method for detecting flavor substances in mutton sheep muscle based on mass spectrometry according to claim 1, characterized in that, The liquid chromatography conditions for the liquid chromatography - mass spectrometry (LC - MS) detection are as follows: the chromatographic column is a C18 reversed - phase chromatographic column (250 mm × 4.6 mm, 5 μm); mobile phase A is 0.1% formic acid aqueous solution, and mobile phase B is acetonitrile; gradient elution program: 0 - 5 minutes, 5% mobile phase B; 5 - 20 minutes, 5% - 95% mobile phase B; 20 - 25 minutes, 95% mobile phase B; 25 - 30 minutes, 95% - 5% mobile phase B; the flow rate is 1 mL / min; the column temperature is 30 °C.

8. The method for detecting flavor substances in mutton sheep muscle based on mass spectrometry according to claim 1, characterized in that, The mass spectrometry conditions for the liquid chromatography - mass spectrometry (LC - MS) detection are as follows: the ion source is an ESI source, in positive ion mode; the spray voltage is 3.5 kV; the capillary temperature is 320 °C; the scanning range is m / z 100 - 1000.

9. The method for detecting flavor substances in mutton sheep muscle based on mass spectrometry according to claim 1, wherein, In the liquid chromatography - mass spectrometry (LC - MS) detection, 10 μL of the sample solution to be tested is injected into the liquid chromatograph. The non - volatile flavor substances are qualitatively analyzed by comparing the retention time and mass spectrum with the standard product, and the non - volatile flavor substances are quantitatively analyzed by the internal standard method.

10. The method for detecting flavor substances in mutton sheep muscle based on mass spectrometry according to claim 1, wherein, In the sample collection and pretreatment steps, the frozen muscle samples are stored in a - 80 °C refrigerator for a long time.

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