Method for identifying tricholoma matsutake and similar edible mushrooms thereof

Abstract

The invention discloses a method for identifying tricholoma matsutake and similar edible mushrooms thereof. The method comprises the following steps: extracting lipid of tricholoma matsutake and similar edible mushrooms thereof; the method comprises the following steps: preparing a sample solution to be tested, analyzing and identifying a lipid molecular structure through ultra-high performance liquid chromatography-high resolution mass spectrometry, and constructing a standard curve through lipid standard substances of different lipid types, so as to realize quantitative analysis of lipid molecules in tricholoma matsutake and similar edible mushrooms. According to the method, qualitative and quantitative results of lipid analysis are synthesized, differential lipid molecules are screened out through least partial square method analysis and p value calculation, and the tricholoma matsutake and similar edible mushroom varieties are identified according to the differential lipid molecules or the differential lipid molecules and the content of the differential lipid molecules. According to the method, only a small amount of edible mushroom powder needs to be taken to extract lipid, the UPLC-HRMS technology is adopted, lipid molecular structure identification and accurate quantification are achieved, and by combining the analysis means of chemometrics, differential lipid can be screened out to be used for identifying tricholoma matsutake and similar edible mushroom varieties.

Description

Technical Field

[0001] The present invention belongs to the field of identification of fungi, and relates to a method for identifying Tricholoma matsutake and its similar edible fungi. Specifically, it relates to obtaining the lipid profiles of Tricholoma matsutake and its similar edible fungi through lipidomics analysis methods, finding differential lipid molecules of different species of edible fungi, and identifying different edible fungi. Background Art

[0002] Tricholoma matsutake, also known as Tricholoma matsutake (S. Ito & S. Imai) Singer, belongs to the subgenus Tricholoma of the family Tricholomataceae. It is mainly distributed in Yunnan, Jilin and other places. Its mushroom cap is flat hemispherical to nearly flat, emitting a strong special mushroom fragrance. It is named because it grows under pine trees and its mushroom is similar to pilose antler. Tricholoma matsutake is mainly for food and can also be used as medicine. It has functions such as strengthening the body, relieving pain, benefiting the stomach and intestines, regulating qi and resolving phlegm, and has the effects of treating diabetes, anti-cancer and preventing premature aging.

[0003] Under the condition of global warming, wild edible mushroom resources are threatened, and the suitable habitats of Tricholoma matsutake have decreased sharply. Although scientific researchers have made a lot of attempts, artificial cultivation has not been fully realized. Due to limited resources, Tricholoma matsutake has been listed as a national key protected wild plant. In addition, due to its unique nutritional and edible value, Tricholoma matsutake has attracted great attention from consumers at home and abroad. Yunnan Province, China exported 174,967 tons of Tricholoma matsutake and its related products in 2022. Therefore, the scarcity of production has driven up the market price of Tricholoma matsutake.

[0004] In recent years, the problem of food adulteration has attracted more and more attention, especially the disguising of inferior products as high-quality products and misleading labels. This problem is particularly common in high-demand products such as Tricholoma matsutake. Many illegal vendors profit by replacing Tricholoma matsutake with visually similar but cheaper and lower-quality fungi such as Stropharia rugosoannulata, Tricholoma sp., Agaricus campestris, Catathelasma laorentou, etc.

[0005] At present, the traditional identification methods of edible fungi are mainly based on appearance, anatomical characteristics, microscopic structure, growth characteristics. However, due to the morphological similarity between different species, these methods often lack accuracy. In contrast, modern technologies such as multiplex real-time PCR, quantitative near-infrared spectroscopy, and DNA barcoding technology provide more reliable identifications, but they are complex, expensive and time-consuming. Therefore, in order to distinguish Tricholoma matsutake from its adulterants, a novel and accurate identification method must be developed. Summary of the Invention

[0006] The object of the present invention is to provide a method for identifying matsutake and its similar edible fungi in view of the unreliability of identifying edible fungi by appearance shape. By using a small amount of samples for lipid extraction and analysis, and integrating the advantages of rapid separation of ultra-high performance liquid chromatography (UPLC) and the high precision and powerful data acquisition ability of high-resolution mass spectrometry (HRMS), the identification of edible fungi varieties is realized.

[0007] The object of the present invention is achieved by the following technical solutions:

[0008] A method for identifying matsutake and its similar edible fungi, comprising: extracting lipids of matsutake and its similar edible fungi; preparing a test sample solution, analyzing and identifying the lipid molecular structure by ultra-high performance liquid chromatography-high resolution mass spectrometry (UPLC-HRMS), and constructing a standard curve through lipid standards of different lipid types to achieve quantitative analysis of lipid molecules in matsutake and its similar edible fungi; comprehensively analyzing the qualitative and quantitative results of lipid analysis, screening out differential lipid molecules through partial least-squares discrimination analysis (PLS-DA) and p-value calculation, and realizing the identification of the types of matsutake and its similar edible fungi according to the differential lipid molecules or the content of differential lipid molecules and differential lipid molecules.

[0009] The similar edible fungi of the matsutake include Stropharia rugosoannulata, Tricholoma sp, Agaricus campestris, and Catathelasma laorentou.

[0010] The method for extracting lipids of matsutake or its similar edible fungi includes: drying and grinding fresh matsutake or its similar edible fungi into powder to obtain matsutake or its similar edible fungi powder; using a mixed solvent with a volume ratio of dichloromethane to methanol of 1:1 to 3:1 as the extraction reagent, vortex-mixing the matsutake or its similar edible fungi powder and the extraction solvent according to the dosage ratio of matsutake or its similar edible fungi powder to the extraction solvent of 1:10 g / mL, performing ultrasonic-assisted extraction at a temperature of 18-22°C for 20-40 min, centrifuging at a temperature of 4°C, taking 200-400 μL of the supernatant, and drying it with nitrogen to obtain lipids of matsutake or its similar edible fungi.

[0011] The traditional Folch method uses chloroform and a mixed solvent of chloroform and methanol as lipid extraction solvents. Although it has a high extraction rate for lipids, it may lead to the co-extraction of some non-lipid components, thus increasing the difficulty of subsequent purification. At the same time, chloroform is carcinogenic and causes relatively large pollution to the environment. Good ventilation is required during experimental operations, and appropriate protective equipment needs to be worn. Compared with the traditional Folch method, dichloromethane has lower toxicity and volatility, and causes less harm to humans and the environment. Due to its relatively low polarity, it co-extracts fewer non-lipid components during the extraction process, thereby improving the purity of lipid extraction.

[0012] Preferably, the extraction reagent is a mixed solvent of dichloromethane and methanol with a volume ratio of 2:1.

[0013] The power of the ultrasonic extraction is 80 Hz.

[0014] Preferably, the temperature of the ultrasonic extraction is 20 °C.

[0015] Preferably, the ultrasonic extraction time is 30 min.

[0016] Preferably, the rotation speed of the centrifugation is 15000 rpm, and the centrifugation time is 10 min.

[0017] Preferably, the volume of the supernatant is 300 μL.

[0018] Preparation of the test sample solution: Add the lipids of Tricholoma matsutake or its similar edible fungi corresponding to 200 - 400 μL of the supernatant to 300 μL of the mobile phase, assist dissolution by ultrasonic waves, and filter through a 0.22 μm nylon hydrophobic filter to obtain the test sample solution.

[0019] Preferably, for the preparation of the test sample solution: Add the lipids of Tricholoma matsutake or its similar edible fungi corresponding to 300 μL of the supernatant to 300 μL of the mobile phase, assist dissolution by ultrasonic waves, and filter through a 0.22 μm nylon hydrophobic filter to obtain the test sample solution.

[0020] The C18 chromatographic column is one of the most commonly used reversed-phase chromatographic columns. Its stationary phase has stable chemical properties and can withstand a wide pH range (conventional pH 2 - 8). The separation effect can be optimized by adjusting parameters such as the composition, flow rate, and temperature of the mobile phase. It is suitable for the separation and analysis of various lipids, including phospholipids, triglycerides, cholesterol, etc., and can provide good separation effects for lipids with different polarities. The C18 chromatographic column has high column efficiency, good reproducibility, and low maintenance costs, making it suitable for long-term use. The HILIC chromatographic column is mainly used for separating polar compounds and has a poorer separation effect for non-polar lipids (such as triglycerides) than the C18 chromatographic column. Moreover, under high pH conditions, its chemical stability is relatively low, and it needs to be used under specific mobile phase conditions, such as high-concentration organic solvents and low-concentration aqueous phases, which limits its operational flexibility. Due to its sensitivity to mobile phase conditions, the maintenance cost of the HILIC chromatographic column is relatively high. The lipids in matsutake and its similar edible fungi are mainly non-polar lipids such as glycerides and glycerophospholipids. Therefore, the present invention uses a C18 chromatographic column as the chromatographic column for ultra-high performance liquid chromatography.

[0021] The conditions of the ultra-high performance liquid chromatography (UPLC) are as follows: Phenomenex Kinetex C18 core-shell universal chromatographic column (2.6 μm, 100×2.1 mm); column temperature: 40 °C; flow rate: 0.2 mL / min; injection volume: 2 μL; Phase A: an ammonium acetate solution with a concentration of 5 mmol / L prepared from a mixed solvent of water:methanol:acetonitrile with a volume ratio of 1:1:1 (water:methanol:acetonitrile (1:1:1, V / V / V) + 5 mM ammonium acetate), Phase B: an ammonium acetate solution with a concentration of 5 mmol / L prepared from a mixed solvent of isopropanol:acetonitrile with a volume ratio of 5:1 (isopropanol:acetonitrile (5:1, V / V) + 5 mM ammonium acetate); the gradient elution conditions are as follows:

[0022]

[0023] The conditions of the high-resolution mass spectrometry (HRMS) are as follows: separate scans in positive and negative ion modes, MS scan range: m / z 120 - 1800 Da, MS / MS scan range: m / z 120 - 1800 Da, the resolutions of the full-scan spectrum and the fragment spectrum are 140000 and 70000 respectively. ESI source parameter settings: spray voltage 3200 V, capillary temperature: 325 °C, heater temperature: 350 °C, collision energy: 25 / 30 / 35 eV, sheath gas flow rate: 40 arbs (high-purity N2, purity > 99%), auxiliary gas flow rate: 20 arbs (high-purity N2, purity > 99%).

[0024] Automatically collect and combine the secondary mass spectra of lipid metabolites contained in Tricholoma matsutake and its similar edible fungi by using the data-dependent acquisition mode of an ultra-high performance liquid chromatography / quadrupole-orbitrap high-resolution mass spectrometer. Use the qualitative software MS-DIAL to determine the retention time, m / z, and secondary mass spectra of the measured lipid metabolites in Tricholoma matsutake and its similar edible fungi; compare the theoretical MS / MS spectra with lipid annotations provided in the software with the LipidBlast database, and qualitatively analyze different lipids according to their fragmentation patterns.

[0025] The inventors conducted a preliminary detection and analysis of the total lipids in Tricholoma matsutake and its similar edible fungi to obtain the species information in the total lipid molecules. According to the preliminary identification results of lipid analysis, 9 non-endogenous lipid molecules were selected as lipid standards. The lipid standards were PC(18:0 / 18:1), PE(17:0 / 17:0), PG(17:0 / 17:0), PA(17:0 / 17:0), PI(8:0 / 8:0), PS(17:0 / 17:0), DGTS(16:0 / 16:0), TG(16:0 / 18:0 / 16:0), Cer(d18:1 / 18:0).

[0026] The process of making the standard curve is as follows: Dissolve the lipid standards in methanol to prepare a standard mixture mother liquor, and dilute it with methanol to a series of concentration solutions, and perform ultra-high performance liquid chromatography-high resolution mass spectrometry (UPLC-HRMS) analysis. Use the concentration of lipid molecules as the abscissa and the peak area of each lipid standard as the ordinate to make the standard curve.

[0027]

[0028] Quantitative analysis method for lipid molecules: Calculate the concentration of each lipid molecule according to the integrated peak area of the extracted ion chromatogram of a single lipid molecule (m / z error value is ±5 ppm) and the standard curve of each lipid standard.

[0029] Screening method for differential lipid molecules: Use the type of lipid molecules and the content of each lipid molecule or its corresponding lg value as variables, perform partial least squares analysis to obtain the VIP value of each lipid molecule; through p-value calculation, screen out lipid molecules with VIP>1 and p<0.05, and further screen out 8 key differential lipids with VIP>3 and / or 10 relatively representative lipid molecules as differential lipid molecules for identifying the species of Tricholoma matsutake and its similar edible fungi.

[0030] The differential lipid molecules of Tricholoma matsutake are PA(18:2 / 18:2), PA(16:0 / 18:2), PE(18:2 / 18:2), DG(16:0 / 18:2), TG(18:0 / 18:1 / 18:2), LPC(18:2), LPE(18:2), PC(18:2 / 18:2), PC(24:0 / 18:2), and PC(24:0 / 18:2) is the unique lipid molecule of Tricholoma matsutake; the differential lipid molecules of Tricholoma bakamatsutake are PA(18:2 / 18:2), PA(16:0 / 18:2), PE(18:2 / 18:2), DG(16:0 / 18:2), TG(18:0 / 18:1 / 18:2), LPC(18:2), LPE(18:2), PC(18:2 / 18:2), PC(18:0 / 18:2), and PC(18:0 / 18:2) is the unique lipid molecule of Tricholoma bakamatsutake; the differential lipid molecules of Tricholoma flavovirens are PA(18:2 / 18:2), PA(16:0 / 18:2), PE(18:2 / 18:2), DG(16:0 / 18:2), TG(18:0 / 18:1 / 18:2), LPC(18:2), LPE(18:2), PE(16:0 / 16:1), and PE(16:0 / 16:1) is the unique lipid molecule of Tricholoma flavovirens; the differential lipid molecules of Agaricus blazei Murill are PA(18:2 / 18:2), PA(16:0 / 18:2), PE(18:2 / 18:2), DG(16:0 / 18:2), TG(18:0 / 18:1 / 18:2), LPC(18:2), LPE(18:2), PC(19:1 / 19:1), and PC(19:1 / 19:1) is the unique lipid molecule of Agaricus blazei Murill; the differential lipid molecules of Lactarius volemus are PA(18:2 / 18:2), PA(16:0 / 18:2), PE(18:2 / 18:2), DG(16:0 / 18:2), TG(18:0 / 18:1 / 18:2), LPC(18:2), LPE(18:2); the content of PC(18:2 / 18:2) in Agaricus blazei Murill is about 1.7 - 1.8 times that in Tricholoma matsutake, the content of PC(18:2 / 18:2) in Tricholoma bakamatsutake, Tricholoma flavovirens, and Lactarius volemus is comparable, and the content of PC(18:2 / 18:2) in Tricholoma bakamatsutake, Tricholoma flavovirens, and Lactarius volemus is: Tricholoma bakamatsutake > Tricholoma flavovirens > Lactarius volemus, and the content of PC(18:2 / 18:2) in Tricholoma bakamatsutake, Tricholoma flavovirens, and Lactarius volemus is about 3 / 5 - 4 / 5 of that in Tricholoma matsutake; the content of LPC(18:2) in Tricholoma matsutake, Tricholoma bakamatsutake, Tricholoma flavovirens, Agaricus blazei Murill, and Lactarius volemus is: Tricholoma matsutake > Agaricus blazei Murill > Tricholoma flavovirens > Lactarius volemus > Tricholoma bakamatsutake, the content of LPC(18:2) in Tricholoma matsutake is the highest, about 1.5 - 1.6 times, about 2 times the content of LPC(18:2) in Tricholoma matsutake var. rufum, about 2.2 - 2.25 times the content of LPC(18:2) in Lactarius volemus, about 3.8 - 4 times the content of LPC(18:2) in Stropharia rugoso-annulata; The contents of LPE(18:2) in Tricholoma matsutake, Stropharia rugoso-annulata, Tricholoma matsutake var. rufum, Agaricus blazei Murrill, and Lactarius volemus are: Tricholoma matsutake > Tricholoma matsutake var. rufum > Agaricus blazei Murrill > Lactarius volemus > Stropharia rugoso-annulata. The content of LPE(18:2) in Tricholoma matsutake is the highest, about 2.6 - 2.7 times the content of LPE(18:2) in Tricholoma matsutake var. rufum, about 3.4 - 3.5 times the content of LPE(18:2) in Agaricus blazei Murrill, about 4.8 - 5 times the content of LPE(18:2) in Lactarius volemus, about 37 times the content of LPE(18:2) in Stropharia rugoso-annulata; The content of PA(18:2 / 18:2) in Stropharia rugoso-annulata is about 1.6 - 1.8 times the content of PA(18:2 / 18:2) in Tricholoma matsutake. The contents of PA(18:2 / 18:2) in Tricholoma matsutake, Agaricus blazei Murrill, and Tricholoma matsutake var. rufum are comparable, and the contents of PA(18:2 / 18:2) in Tricholoma matsutake, Agaricus blazei Murrill, and Tricholoma matsutake var. rufum are: Tricholoma matsutake > Agaricus blazei Murrill > Tricholoma matsutake var. rufum. The content of PA(18:2 / 18:2) in Tricholoma matsutake is about 3.6 - 4 times the content of PA(18:2 / 18:2) in Lactarius volemus; The content of PA(16:0 / 18:2) in Tricholoma matsutake is relatively low, about 2 - 2.5 times the content of PA(16:0 / 18:2) in Lactarius volemus, about 1 / 5 - 1 / 4 of the content of PA(16:0 / 18:2) in Stropharia rugoso-annulata, about 1 / 3 - 2 / 5 of the content of PA(16:0 / 18:2) in Agaricus blazei Murrill, about 3 / 4 - 4 / 5 of the content of PA(16:0 / 18:2) in Tricholoma matsutake var. rufum; The content of PE(18:2 / 18:2) in Tricholoma matsutake is the highest, comparable to the content of PE(18:2 / 18:2) in Agaricus blazei Murrill, about 1.2 times the content of PE(18:2 / 18:2) in Agaricus blazei Murrill, about 2.7 - 3 times the content of PE(18:2 / 18:2) in Tricholoma matsutake var. rufum, about 3.9 - 4 times the content of PE(18:2 / 18:2) in Stropharia rugoso-annulata, about 12 - 12.5 times; The content of DG(16:0 / 18:2) in Rhodophypanus nigripes is much higher than that in the other four edible fungi. The contents of DG(16:0 / 18:2) in Rhodophypanus nigripes, Strobilomyces floccopus, Agaricus blazei, Tricholoma matsutake, and Gerronema virgatum are as follows: Rhodophypanus nigripes > Strobilomyces floccopus > Agaricus blazei > Tricholoma matsutake > Gerronema virgatum. The content of DG(16:0 / 18:2) in Strobilomyces floccopus is about 9 / 50 - 1 / 5 times that in Rhodophypanus nigripes. The content of DG(16:0 / 18:2) in Agaricus blazei is about 1 / 10 - 3 / 20 times that in Rhodophypanus nigripes. The content of DG(16:0 / 18:2) in Tricholoma matsutake is about 1 / 11 - 1 / 10 times that in Rhodophypanus nigripes. The content of DG(16:0 / 18:2) in Gerronema virgatum is about 1 / 25 times that in Rhodophypanus nigripes. The contents of TG(18:1 / 18:1 / 18:2) in Rhodophypanus nigripes, Tricholoma matsutake, Strobilomyces floccopus, Agaricus blazei, and Gerronema virgatum are as follows: Strobilomyces floccopus > Tricholoma matsutake > Rhodophypanus nigripes > Gerronema virgatum > Agaricus blazei. The contents of TG(18:1 / 18:1 / 18:2) in Tricholoma matsutake and Rhodophypanus nigripes are both about 2 - 2.2 times that in Agaricus blazei. The content of TG(18:1 / 18:1 / 18:2) in Gerronema virgatum is about 1.5 - 1.6 times that in Agaricus blazei. The content of TG(18:1 / 18:1 / 18:2) in Strobilomyces floccopus is about 3.6 - 3.65 times that in Tricholoma matsutake.

[0031] Furthermore, DGTS(17:0 / 18:2) and PE(17:0 / 18:2) were only detected in Tricholoma matsutake and Strobilomyces floccopus. The content of DGTS(17:0 / 18:2) in Strobilomyces floccopus was about 5.9 - 6 times that in Tricholoma matsutake, and the content of PE(17:0 / 18:2) in Strobilomyces floccopus was about 1.8 - 2 times that in Tricholoma matsutake. PE(24:0 / 18:2) was only detected in Tricholoma matsutake and Agaricus blazei Murrill. The content of PE(24:0 / 18:2) in Agaricus blazei Murrill was about 5 - 5.5 times that in Tricholoma matsutake. Except for Rosenbergiella neofloccosa, PE(18:0 / 18:2) was detected in the other four edible fungi. The content of PE(18:0 / 18:2) in Tricholoma matsutake was the highest, about 2 - 2.2 times that in Strobilomyces floccopus, about 2.9 - 3 times that in Agaricus blazei Murrill, and about 4.8 - 5 times that in Agaricus bitorquis. Except for Rosenbergiella neofloccosa, LPS(18:2) was detected in the other four edible fungi. The content of LPS(18:2) in Tricholoma matsutake was the highest, about 2 - 2.2 times that in Strobilomyces floccopus, about 4.5 times that in Agaricus blazei Murrill, and about 19 - 20 times that in Agaricus bitorquis. The content of LPI(18:2) in Rosenbergiella neofloccosa was greater than that of PI(18:2) in the other four edible fungi. The content of LPI(18:2) in Agaricus bitorquis, Tricholoma matsutake, Strobilomyces floccopus, Agaricus blazei Murrill, and Rosenbergiella neofloccosa was: Rosenbergiella neofloccosa > Strobilomyces floccopus > Tricholoma matsutake > Agaricus blazei Murrill > Agaricus bitorquis. The content of LPI(18:2) in Rosenbergiella neofloccosa was about 1.5 - 1.55 times that in Strobilomyces floccopus, about 1.9 - 2 times that in Tricholoma matsutake, about 2.8 - 2.85 times that in Agaricus blazei Murrill, and about 8 times that in Agaricus bitorquis.

[0032] Furthermore, one relatively representative lipid molecule, PE(18:2 / 17:1; O2), was screened out as a differential lipid molecule for identifying Tricholoma matsutake and its similar edible fungi species. PE(18:2 / 17:1; O2) only exists in Strobilomyces floccopus and Agaricus blazei Murrill, and the content of PE(18:2 / 17:1; O2) in Agaricus blazei Murrill is about 1.5 - 1.6 times that in Strobilomyces floccopus.

[0033] A method for identifying Tricholoma matsutake and its similar edible fungi includes:

[0034] Step (1), extracting the lipids of Tricholoma matsutake and its similar edible fungi;

[0035] Step (2): Prepare the test sample solution, identify the lipid molecular structure by ultra-high performance liquid chromatography-high resolution mass spectrometry (UPLC-HRMS), and construct a standard curve with lipid standards of different lipid types to achieve quantitative analysis of lipid molecules in Tricholoma matsutake and its similar edible fungi. Based on the qualitative and quantitative results of lipid analysis, differential lipid molecules are screened through partial least squares analysis and p-value calculation.

[0036] Step (3): Extract the lipids of Tricholoma matsutake and its similar edible fungi according to step (1), quantify the screened differential lipid molecules according to step (2), and identify the types of Tricholoma matsutake and its similar edible fungi based on the differential lipid molecules or the content of differential lipid molecules and differential lipid molecules.

[0037] Advantages of the present invention:

[0038] 1. The present invention only needs to take a small amount (about 0.15 g) of the powder of Tricholoma matsutake and its similar edible fungi to extract lipids, and uses the UPLC-HRMS technology to achieve high-throughput, high-sensitivity, and high-resolution identification and accurate quantification of lipid molecular structures.

[0039] 2. In the UPLC analysis of the present invention, a C18 chromatographic column (neutral and lipophilic liquid chromatography) is used to separate the lipids of edible fungi. Its stationary phase has stable chemical properties, can tolerate a wide pH range (conventional pH 2-8), has a smoother gradient elution, small baseline fluctuations, and a short equilibration time (5-10 minutes), and can analyze and identify more lipid molecules. At the same time, by analyzing different lipid classes contained in the sample, non-endogenous lipid molecules not contained in the sample are respectively selected as standards to achieve quantitative analysis of different classes of lipids.

[0040] 3. According to the mass spectrometry analysis results of the lipids of Tricholoma matsutake and its similar edible fungi, combined with the analysis methods of chemometrics, differential lipids can be screened for the discrimination of edible fungi species, providing guarantee for the quality control and adulteration identification of some precious edible fungi (such as Tricholoma matsutake) with similar traits after drying, improving the variety identification efficiency, and avoiding adulteration behavior.

[0041] 4. Different from accurately identifying poisonous fungi, the present invention can be widely applied to the adulteration identification of Tricholoma matsutake and its similar edible fungi in the market, greatly guaranteeing the authenticity of products and the interests of the people. Description of the drawings

[0042] Figure 1 : Total ion current chromatograms (TICs) of lipid molecular species of QC samples in the ESI(+)(left) mode and ESI(-)(right) mode.

[0043] Figure 2 : Distribution diagram of the coefficient of variation (CV) of all features of untargeted lipidomics.

[0044] Figure 3 : PCA plot of lipid molecules of 5 edible fungi; in the figure, each point represents an experimental sample, and each color represents an edible fungus.

[0045] Figure 4 : VIP value plot of lipid molecules of 5 edible fungi.

[0046] Figure 5 : Differential lipid heat map of lipid molecules of 5 edible fungi.

[0047] Figure 6 : PLS-DA score plot of lipidomics data of 5 edible fungi based on 50 differential lipids.

[0048] Figure 7 : 200 permutation test plot of the new PLS-DA model.

[0049] Figure 8 : PLS-DA score plot of 5 edible fungi and real samples based on 50 identified differential lipids. Detailed implementation manners

[0050] The technical solutions of the present invention will be further described below through specific implementation manners.

[0051] Example 1

[0052] 5 edible fungi: Tricholoma matsutake, Stropharia rugosoannulata, Tricholoma sp, Agaricus campestris, Catathelasma laorentou, all harvested in Yunnan.

[0053] 1. Lipid extraction

[0054] Fresh edible fungi are dried at 35 °C, ground into powder, and the edible fungus powder is stored at -20 °C.

[0055] Take 0.15 g of edible fungus powder, mix the edible fungus powder with 1.5 mL of a mixed solvent of dichloromethane and methanol (dichloromethane: methanol = 2:1, V / V), vortex for 2 min, ultrasonically extract for 30 min at a temperature of 20 °C and a power of 80 Hz, centrifuge at a temperature of 4 °C and a rotation speed of 15000 rpm for 10 min, take 300 μL of the supernatant, dry it with nitrogen to obtain edible fungus lipids, and store them in a -20 °C refrigerator.

[0056] 2. UPLC-QE Orbitrap / MS / MS analysis and lipid molecule structure identification and quantitative analysis

[0057] Preparation of the test sample solution: The edible mushroom lipid obtained by drying 300 μL of the supernatant was added to 300 μL of the mobile phase, sonicated for dissolution, and filtered through a 0.22 μm nylon hydrophobic filter to obtain the test sample solution.

[0058] Analysis was performed by ultra-high performance liquid chromatography-high resolution mass spectrometry (UPLC-HRMS).

[0059] Ultra-high performance liquid chromatography (UPLC) conditions: Phenomenex Kinetex C18 core-shell universal chromatographic column (2.6 μm, 100×2.1 mm); column temperature: 40 °C; flow rate: 0.2 mL / min; injection volume: 2 μL; phase A: water: methanol: acetonitrile (1:1:1, V / V / V) + 5 mM ammonium acetate, phase B: isopropanol: acetonitrile (5:1, V / V) + 5 mM ammonium acetate; the gradient elution conditions are shown in Table 1.

[0060] Table 1: Ultra-high performance liquid chromatography gradient elution conditions

[0061]

[0062] Mass spectrometry (HRMS) conditions: Scanning mode: separate scanning in positive and negative ion modes, MS scanning range: m / z 120 - 1800 Da, MS / MS scanning range: m / z 120 - 1800 Da, the resolutions of the full scan spectrum and the fragment spectrum are 140000 and 70000 respectively. ESI source parameter settings: spray voltage 3200 V, capillary temperature: 325 °C, heater temperature: 350 °C, collision energy: 25 / 30 / 35 eV, sheath gas flow rate: 40 arbs (high-purity N2, purity > 99%), auxiliary gas flow rate: 20 arbs (high-purity N2, purity > 99%).

[0063] Data processing: The analysis of the original data was completed using software such as MS-DIAL ver.4.92, SIMCA 14.1, SPSS26.0, ChemDraw 14.0, and Origin2021.

[0064] In high-resolution mass spectrometry, ionized lipid molecules reach the collision cell through a quadrupole mass analyzer. Under collision-induced dissociation conditions, chemical bonds with lower bond energy in the molecular structure break, forming secondary ion fragments. The data-dependent acquisition mode of an ultra-high performance liquid chromatography / quadrupole-orbitrap high-resolution mass spectrometer is used to automatically collect and combine the secondary mass spectra of lipid metabolites contained in each edible mushroom. The original data file (.raw) obtained from the mass spectrometry is converted into the Analysis Baes File format (.abf) using the AnalysisBaseFileConverter.exe software, and then the data is imported into the MS-DIAL software for analysis. The retention time, m / z, and secondary mass spectra of the lipid metabolites measured in each edible mushroom are determined using the MS-DIAL software. Different lipids are qualitatively analyzed based on the theoretical MS / MS spectra with lipid annotations provided in the software and compared with the LipidBlast database, and the specific structure of the lipid molecules is deduced.

[0065] The results showed that a total of 309 lipid molecules were detected in 5 edible mushrooms, mainly including glycerides, glycerophospholipids, and sphingolipids. More lipid molecules were detected in Tricholoma matsutake, Tricholoma lobayense, and Tricholoma mongolicum, with 165, 168, and 166 species respectively, followed by Agaricus blazei with 135 lipid molecules, and the fewest lipid molecules were detected in Gerronema virgatum, with 91 species. There were significant differences in the lipid types contained in different edible mushrooms. A total of 42 lipid molecules were commonly detected in the five edible mushrooms, and the most common lipid molecules were detected in Tricholoma lobayense, with 42 species. This proved that there were significant differences in the lipid composition between Tricholoma lobayense and other types of edible mushrooms. At the same time, the fewest lipid molecules were detected in Gerronema virgatum, and the fewest unique lipid molecules were also detected, with 13 species.

[0066] Preparation of the standard curve: Nine non-endogenous lipid molecules were selected as lipid standards, namely PC(18:0 / 18:1), PE(17:0 / 17:0), PG(17:0 / 17:0), PA(17:0 / 17:0), PI(8:0 / 8:0), PS(17:0 / 17:0), DGTS(16:0 / 16:0), TG(16:0 / 18:0 / 16:0), and Cer(d18:1 / 18:0); The nine lipid standards were dissolved in methanol to prepare a standard mixture stock solution, and then diluted with methanol to a series of concentration solutions (10 - 5000 ng / ml) for ultra-high performance liquid chromatography-high resolution mass spectrometry (UPLC-HRMS) analysis. The concentration of the lipid molecules was used as the abscissa, and the peak area of each type of lipid standard was used as the ordinate to prepare the standard curve (Table 2). Based on the integrated peak area of the extracted ion chromatogram of a single lipid molecule (m / z error value of ±5 ppm), the concentration of each lipid molecule was calculated according to the standard curve of each lipid standard.

[0067] Table 2: Standard Curve of Lipid Standards

[0068]

[0069] Note: x represents the concentration of lipid molecules (mg / mL), and y represents the peak area of lipids.

[0070] 3. Quality Control (QC)

[0071] QC samples are an essential part of the sample detection process and can be used to study the stability and repeatability of the method. In this experiment, the QC preparation was completed by equally mixing the samples tested for each edible mushroom. In the entire run batch, one QC sample was injected into every four experimental samples. To ensure the reliability of the results, after all sample detections were completed, the original lipid profile data of the QC samples were collected for data preprocessing to obtain a series of characteristic ion peaks, the average value of the peak area was calculated, and its standard deviation (SD) was obtained. Finally, the coefficient of variation (CV) value was calculated. Samples of the same batch were continuously analyzed on the instrument.

[0072] 4. Chemometric Analysis

[0073] Combined with the qualitative and quantitative results of lipids by MSDIAL ver.4.92, SIMCA 14.1 was used to analyze the detection results with a mathematical model.

[0074] Total ion chromatograms (TIC) of 5 edible mushroom lipid samples in positive ion (left) and negative ion (right) modes are as Figure 1 shown. The peak intensities and retention times of these edible mushroom lipid samples are consistent, and the TIC diagrams almost completely overlap, demonstrating the high signal stability of the mass spectrometer. In addition, the CV values of more than 90% of the detected features are ≤10%, and 97.8% of the CV values are ≤20%( Figure 2 ), confirming the stability and reliability of the analysis system.

[0075] Principal component analysis (PCA) was performed on the types and contents of lipid molecules of 5 edible mushrooms to show the differences between the lipid components of 5 edible mushrooms. The PCA score plots of the five groups of samples are as Figure 3 shown, R 2 Xcum = 0.995, Q 2 cum = 0.989, and the cumulative contribution rate of the two principal components reaches 0.667, indicating that the PCA model has a good fitting effect. The separation and clustering trends of different types of samples are obvious, indicating that their lipids have obvious differences. The results show that: through PCA analysis, better analysis results can be obtained, that is, samples within the group are aggregated and samples between groups are dispersed, and the experimental data meet the conditions of PCA analysis.

[0076] Taking the lipid molecular types and the contents of each lipid molecule as variables, partial least squares discrimination analysis (PLS-DA) was performed to obtain the VIP values of each lipid molecule ( Figure 4 ). Lipids with VIP>1 were regarded as key lipids. Further, through p-value calculation, 50 differential lipid molecules with VIP>1 and p<0.05 were screened out, and a heat map of the contents was drawn, as Figure 5 shown. The VIP values of PC(18:2 / 18:2), LPC(18:2), and LPE(18:2) were higher than those of other lipid molecules (VIP>4). Therefore, they contributed greatly to the identification of these 5 edible fungi.

[0077] As shown in Table 3, the contents and VIP values of 50 differential lipids of 5 edible fungi were selected, and 8 key differential lipids with VIP>3 and 11 relatively representative lipid molecules were analyzed.

[0078] Table 3: Contents, VIP values, and P values of 50 differential lipids of 5 edible fungi

[0079]

[0080]

[0081]

[0082] Note: nd indicates not detected.

[0083] As shown in Table 3, among these differential lipids, the VIP values of PA(18:2 / 18:2), PA(16:0 / 18:2), PE(18:2 / 18:2), DG(16:0 / 18:2), and TG(18:0 / 18:1 / 18:2) were greater than 3 and they were all distributed among the 5 edible fungi.

[0084] The differential lipid molecules of Tricholoma matsutake are PA(18:2 / 18:2), PA(16:0 / 18:2), PE(18:2 / 18:2), DG(16:0 / 18:2), TG(18:0 / 18:1 / 18:2), LPC(18:2), LPE(18:2), PC(18:2 / 18:2), PC(24:0 / 18:2), and PC(24:0 / 18:2) is the unique lipid molecule of Tricholoma matsutake; the differential lipid molecules of Tricholoma akahatsu are PA(18:2 / 18:2), PA(16:0 / 18:2), PE(18:2 / 18:2), DG(16:0 / 18:2), TG(18:0 / 18:1 / 18:2), LPC(18:2), LPE(18:2), PC(18:2 / 18:2), PC(18:0 / 18:2), and PC(18:0 / 18:2) is the unique lipid molecule of Tricholoma akahatsu; the differential lipid molecules of Tricholoma bakamatsutake are PA(18:2 / 18:2), PA(16:0 / 18:2), PE(18:2 / 18:2), DG(16:0 / 18:2), TG(18:0 / 18:1 / 18:2), LPC(18:2), LPE(18:2), PE(16:0 / 16:1), and PE(16:0 / 16:1) is the unique lipid molecule of Tricholoma bakamatsutake; the differential lipid molecules of Agaricus blazei are PA(18:2 / 18:2), PA(16:0 / 18:2), PE(18:2 / 18:2), DG(16:0 / 18:2), TG(18:0 / 18:1 / 18:2), LPC(18:2), LPE(18:2), PC(19:1 / 19:1), and PC(19:1 / 19:1) is the unique lipid molecule of Agaricus blazei; the differential lipid molecules of Strobilomyces strobilaceus are PA(18:2 / 18:2), PA(16:0 / 18:2), PE(18:2 / 18:2), DG(16:0 / 18:2), TG(18:0 / 18:1 / 18:2), LPC(18:2), LPE(18:2); the content of PC(18:2 / 18:2) in Agaricus blazei is about 1.7 times that in Tricholoma matsutake, the content of PC(18:2 / 18:2) in Tricholoma akahatsu, Tricholoma bakamatsutake, and Strobilomyces strobilaceus is comparable, and the content of PC(18:2 / 18:2) in Tricholoma akahatsu, Tricholoma bakamatsutake, and Strobilomyces strobilaceus is: Tricholoma akahatsu > Tricholoma bakamatsutake > Strobilomyces strobilaceus, and the content of PC(18:2 / 18:2) in Tricholoma akahatsu, Tricholoma bakamatsutake, and Strobilomyces strobilaceus is about 3 / 4, 2 / 3, and 3 / 5 of the content of PC(18:2 / 18:2) in Tricholoma matsutake respectively; the content of LPC(18:2) in Tricholoma matsutake, Tricholoma akahatsu, Tricholoma bakamatsutake, Agaricus blazei, and Strobilomyces strobilaceus is: Tricholoma matsutake > Agaricus blazei > Tricholoma bakamatsutake > Strobilomyces strobilaceus > Tricholoma akahatsu, the content of LPC(18:2) in Tricholoma matsutake is the highest, about 1.5 times, about 2 times the content of LPC(18:2) in Tricholoma matsutake var. albuminosum, about 2 times the content of LPC(18:2) in Lactarius volemus, and about 4 times the content of LPC(18:2) in Tricholoma bakamatsutake; The contents of LPE(18:2) in Tricholoma matsutake, Tricholoma bakamatsutake, Tricholoma matsutake var. albuminosum, Agaricus blazei, and Lactarius volemus are: Tricholoma matsutake > Tricholoma matsutake var. albuminosum > Agaricus blazei > Lactarius volemus > Tricholoma bakamatsutake. The content of LPE(18:2) in Tricholoma matsutake is the highest, about 2.7 times the content of LPE(18:2) in Tricholoma matsutake var. albuminosum, about 3.4 times the content of LPE(18:2) in Agaricus blazei, about 4.8 times the content of LPE(18:2) in Lactarius volemus, and about 37 times the content of LPE(18:2) in Tricholoma bakamatsutake; The content of PA(18:2 / 18:2) in Tricholoma bakamatsutake is about 1.6 times the content of PA(18:2 / 18:2) in Tricholoma matsutake. The contents of PA(18:2 / 18:2) in Tricholoma matsutake, Agaricus blazei, and Tricholoma matsutake var. albuminosum are comparable, and the contents of PA(18:2 / 18:2) in Tricholoma matsutake, Agaricus blazei, and Tricholoma matsutake var. albuminosum are: Tricholoma matsutake > Agaricus blazei > Tricholoma matsutake var. albuminosum. The content of PA(18:2 / 18:2) in Tricholoma matsutake is about 4 times the content of PA(18:2 / 18:2) in Lactarius volemus; The content of PA(16:0 / 18:2) in Tricholoma matsutake is relatively low, about 2 times the content of PA(16:0 / 18:2) in Lactarius volemus, about 1 / 5 of the content of PA(16:0 / 18:2) in Tricholoma bakamatsutake, about 0.38 of the content of PA(16:0 / 18:2) in Agaricus blazei, and about 0.77 of the content of PA(16:0 / 18:2) in Tricholoma matsutake var. albuminosum; The content of PE(18:2 / 18:2) in Tricholoma matsutake is the highest, comparable to the content of PE(18:2 / 18:2) in Agaricus blazei, about 1.2 times the content of PE(18:2 / 18:2) in Agaricus blazei, about 2.7 times the content of PE(18:2 / 18:2) in Tricholoma matsutake var. albuminosum, about 4 times the content of PE(18:2 / 18:2) in Tricholoma bakamatsutake, and about 12 times the content of PE(18:2 / 18:2) in Lactarius volemus; The content of DG(16:0 / 18:2) in Tricholoma bakamatsutake is much higher than that of the other four edible fungi. The contents of DG(16:0 / 18:2) in Tricholoma bakamatsutake, Tricholoma matsutake var. albuminosum, Agaricus blazei, Tricholoma matsutake, and Lactarius volemus are: Tricholoma bakamatsutake > Tricholoma matsutake var. albuminosum > Agaricus blazei > Tricholoma matsutake > Lactarius volemus. The content of DG(16:0 / 18:2) in Tricholoma matsutake var. albuminosum is about 0.19 times that of Tricholoma bakamatsutake, the content of DG(16:0 / 18:2) in Agaricus blazei is about 0.14 times that of Tricholoma bakamatsutake, the content of DG(16:0 / 18:2) in Tricholoma matsutake is about 1 / 11 times that of Tricholoma bakamatsutake, and the content of DG(16:0 / 18:2) in Lactarius volemus is about 1 / 25 of that of Tricholoma bakamatsutake; The contents of TG(18:1 / 18:1 / 18:2) in Tricholoma bakamatsutake, Tricholoma matsutake, Tricholoma matsutake var. albuminosum, Agaricus blazei, and Lactarius volemus are: Tricholoma matsutake var. albuminosum > Tricholoma matsutake > Tricholoma bakamatsutake > Lactarius volemus > Agaricus blazei. The contents of TG(18:1 / 18:1 / 18:2) in Tricholoma matsutake and Tricholoma bakamatsutake are both about 2 - 2 times the content in Agaricus blazei.The content of TG(18:1 / 18:1 / 18:2) in the old man's head is about 2 times that in Agaricus blazei, the content of TG(18:1 / 18:1 / 18:2) in Flammulina velutipes is about 1.6 times that in Agaricus blazei, and the content of TG(18:1 / 18:1 / 18:2) in Anthracophyllum discolor is about 3.6 times that in Tricholoma matsutake.

[0085] DGTS(17:0 / 18:2) and PE(17:0 / 18:2) were only detected in Tricholoma matsutake and Anthracophyllum discolor. The content of DGTS(17:0 / 18:2) in Anthracophyllum discolor is about 5.9 times that in Tricholoma matsutake, and the content of PE(17:0 / 18:2) in Anthracophyllum discolor is about 1.88 times that in Tricholoma matsutake; PE(24:0 / 18:2) was only detected in Tricholoma matsutake and Agaricus blazei, and the content of PE(24:0 / 18:2) in Agaricus blazei is about 5.3 times that in Tricholoma matsutake; Except for the old man's head, PE(18:0 / 18:2) was detected in the other four edible fungi. The content of PE(18:0 / 18:2) in Tricholoma matsutake is the highest, about 2.2 times that in Anthracophyllum discolor, about 2.9 times that in Agaricus blazei, and about 4.8 times that in Stropharia rugosoannulata; Except for the old man's head, LPS(18:2) was detected in the other four edible fungi. The content of LPS(18:2) in Tricholoma matsutake is the highest, about 2.1 times that in Anthracophyllum discolor, about 4.5 times that in Agaricus blazei, and about 19.4 times that in Stropharia rugosoannulata; The content of LPI(18:2) in the old man's head is greater than that of PI(18:2) in the other four edible fungi. The content of LPI(18:2) in Stropharia rugosoannulata, Tricholoma matsutake, Anthracophyllum discolor, Agaricus blazei, and the old man's head is: the old man's head > Anthracophyllum discolor > Tricholoma matsutake > Agaricus blazei > Stropharia rugosoannulata. The content of LPI(18:2) in the old man's head is about 1.5 times that in Anthracophyllum discolor, about 1.9 times that in Tricholoma matsutake, about 2.8 times that in Agaricus blazei, and about 8 times that in Stropharia rugosoannulata.

[0086] PE(18:2 / 17:1; O2) only exists in Anthracophyllum discolor and Agaricus blazei, and the content of PE(18:2 / 17:1; O2) in Agaricus blazei is about 1.56 times that in Anthracophyllum discolor.

[0087] The identification of Tricholoma matsutake and its similar edible fungi can be achieved according to the differential lipid molecules or the content of differential lipid molecules and differential lipid molecules.

[0088] Example 2

[0089] The steps of lipid extraction, UPLC-QE Orbitrap / MS / MS analysis, lipid molecular structure identification and quantitative analysis, and chemometric analysis refer to Example 1.

[0090] To evaluate the ability of differential lipid identification and prediction of Tricholoma matsutake and its similar edible mushrooms, the inventors selected 40 additional mushroom samples for verification, including 6 samples of Tricholoma lobayense, 9 samples of Tricholoma matsutake, 9 samples of Tricholoma bakamatsutake, 10 samples of Agaricus blazei, and 6 samples of Gerronema virgatum. First, a PLS-DA model was reconstructed using 50 differential lipids with VIP>1 and p<0.05 screened in Example 1, and then 200 permutation tests were performed to confirm the stability of the model ( Figure 6 and Figure 7 ). The new model was as effective as the original model in distinguishing samples from five different edible mushrooms. The evaluation parameters (R 2 X(cum)=0.997, R 2 (cum)=0.998 and Q 2 (cum)=0.996) showed excellent model fitting and prediction capabilities. In addition, the results of 200 permutation tests strongly indicated that the new model was effective because the blue Q 2 values on the left and the green R 2 values were uniformly lower compared to the original points on the right.

[0091] Finally, an external validation test was conducted, and 40 additional edible mushroom samples from 5 varieties were blindly tested and labeled as unknown. These samples were imported into the PLS-DA prediction model to predict their species without prior classification. As shown in Table 4, the overall identification accuracy rate was 92.5%, and all blind test samples of all species were accurately classified except for 3 Agaricus blazei samples. In addition, the distribution of most test samples corresponded to their respective standard references, which was also consistent with the prediction of the algorithm, except for Test 4 (Agaricus blazei)( Figure 8 ). The results of the external validation further demonstrated the practicality of the new PLS-DA model.

[0092] Table 4. External validation results of the partial least squares discriminant analysis (PLS-DA) model reconstructed based on key differential lipids

[0093]

Claims

1. A method for identifying matsutake and its similar edible fungi, characterized in that: Including: Extracting the lipids of Tricholoma matsutake and its similar edible fungi; Preparing a test sample solution, analyzing and identifying the lipid molecular structure by ultra-high performance liquid chromatography-high resolution mass spectrometry, and constructing a standard curve through lipid standards of different lipid types to achieve quantitative analysis of lipid molecules in Tricholoma matsutake and its similar edible fungi; comprehensively based on the qualitative and quantitative results of lipid analysis, screening out differential lipid molecules through partial least squares analysis and p-value calculation, and identifying the types of Tricholoma matsutake and its similar edible fungi according to the differential lipid molecules or the content of differential lipid molecules and differential lipid molecules.

2. The identification method of Tricholoma matsutake and its similar edible fungi according to claim 1, characterized in that: The similar edible fungi of Tricholoma matsutake described are Tricholoma lobayense, Tricholoma bakamatsutake, Agaricus blazei Murill, and Gerocomellus candidus.

3. The identification method of Tricholoma matsutake and its similar edible fungi according to claim 1, characterized in that: The method for extracting the lipids of Tricholoma matsutake or its similar edible fungi includes: drying and grinding fresh Tricholoma matsutake or its similar edible fungi into powder to obtain Tricholoma matsutake or its similar edible fungi powder; using a mixed solvent with a volume ratio of dichloromethane to methanol of 1:1 to 3:1 as the extraction reagent, vortex-mixing the Tricholoma matsutake or its similar edible fungi powder and the extraction solvent according to a dosage ratio of 1:10 g / mL of Tricholoma matsutake or its similar edible fungi powder to the extraction solvent, performing ultrasonic-assisted extraction at a temperature of 18-22 °C for 20-40 min, centrifuging at a temperature of 4 °C, taking 200-400 μL of the supernatant, and drying it with nitrogen to obtain the lipids of Tricholoma matsutake or its similar edible fungi.

4. The identification method of Tricholoma matsutake and its similar edible fungi according to claim 1 or 3, characterized in that: Preparation of the test sample solution: Adding the lipids of Tricholoma matsutake or its similar edible fungi corresponding to 200-400 μL of the supernatant into 300 μL of the mobile phase, assisting dissolution by ultrasound, and filtering with a 0.22 μm nylon hydrophobic filter to obtain the test sample solution.

5. The identification method of Tricholoma matsutake and its similar edible fungi according to claim 1, characterized in that: The ultra-high performance liquid chromatography conditions: Phenomenex Kinetex C18 core-shell universal chromatographic column (2.6 μm, 100×2.1 mm); column temperature: 40 °C; flow rate: 0.2 mL / min; injection volume: 2 μL; Phase A: an ammonium acetate solution with a concentration of 5 mmol / L prepared from a mixed solvent with a volume ratio of water:methanol:acetonitrile of 1:1:1, Phase B: an ammonium acetate solution with a concentration of 5 mmol / L prepared from a volume ratio of isopropanol:acetonitrile of 5:1; the gradient elution conditions are as follows: The high resolution mass spectrometry conditions: Scanning separately in positive and negative ion modes, MS scanning range: m / z 120-1800 Da, MS / MS scanning range: m / z 120-1800 Da, the resolutions of the full scan spectrum and the fragment spectrum are respectively: 140000, 70000; ESI source parameter settings: spray voltage 3200 V, capillary temperature: 325 °C, heater temperature: 350 °C, collision energy: 25 / 30 / 35 eV, sheath gas is N2 with a purity > 99%, flow rate: 40 arbs, auxiliary gas is N2 with a purity > 99%, flow rate: 20 arbs.

6. The identification method of Tricholoma matsutake and its similar edible fungi according to claim 1, characterized in that: The lipid standards are PC(18:0 / 18:1), PE(17:0 / 17:0), PG(17:0 / 17:0), PA(17:0 / 17:0), PI(8:0 / 8:0), PS(17:0 / 17:0), DGTS(16:0 / 16:0), TG(16:0 / 18:0 / 16:0), Cer(d18:1 / 18:0).

7. The identification method of Tricholoma matsutake and its similar edible fungi according to claim 1, characterized in that: Screening method for differential lipid molecules: Taking the type of lipid molecules and the content of each lipid molecule or its corresponding lg value as variables, performing partial least squares analysis to obtain the VIP value of each lipid molecule; Through p-value calculation, screening out lipid molecules with VIP>1 and p<0.05, and further screening out 8 key differential lipids with VIP>3 and / or 10 relatively representative lipid molecules as differential lipid molecules for identifying matsutake and its similar edible mushroom species.

8. The identification method of Tricholoma matsutake and its similar edible fungi according to claim 1 or 7, characterized in that: The differential lipid molecules of matsutake are PA(18:2 / 18:2), PA(16:0 / 18:2), PE(18:2 / 18:2), DG(16:0 / 18:2), TG(18:0 / 18:1 / 18:2), LPC(18:2), LPE(18:2), PC(18:2 / 18:2), PC(24:0 / 18:2), and PC(24:0 / 18:2) is a unique lipid molecule of matsutake; The differential lipid molecules of russula are PA(18:2 / 18:2), PA(16:0 / 18:2), PE(18:2 / 18:2), DG(16:0 / 18:2), TG(18:0 / 18:1 / 18:2), LPC(18:2), LPE(18:2), PC(18:2 / 18:2), PC(18:0 / 18:2), and PC(18:0 / 18:2) is a unique lipid molecule of russula; The differential lipid molecules of Tricholoma matsutakeoides are PA(18:2 / 18:2), PA(16:0 / 18:2), PE(18:2 / 18:2), DG(16:0 / 18:2), TG(18:0 / 18:1 / 18:2), LPC(18:2), LPE(18:2), PE(16:0 / 16:1), and PE(16:0 / 16:1) is a unique lipid molecule of Tricholoma matsutakeoides; The differential lipid molecules of Agaricus blazei are PA(18:2 / 18:2), PA(16:0 / 18:2), PE(18:2 / 18:2), DG(16:0 / 18:2), TG(18:0 / 18:1 / 18:2), LPC(18:2), LPE(18:2), PC(19:1 / 19:1), and PC(19:1 / 19:1) is a unique lipid molecule of Agaricus blazei; The differential lipid molecules of the old man's head mushroom are PA(18:2 / 18:2), PA(16:0 / 18:2), PE(18:2 / 18:2), DG(16:0 / 18:2), TG(18:0 / 18:1 / 18:2), LPC(18:2), LPE(18:2); The content of PC(18:2 / 18:2) in Agaricus blazei is about 1.7 - 1.8 times that of PC(18:2 / 18:2) in Tricholoma matsutake. The content of PC(18:2 / 18:2) is comparable in Russula brevipes, Flammulina velutipes, and the old man's head mushroom, and the content of PC(18:2 / 18:2) in Russula brevipes, Flammulina velutipes, and the old man's head mushroom is: Russula brevipes > Flammulina velutipes > the old man's head mushroom. The content of PC(18:2 / 18:2) in Russula brevipes, Flammulina velutipes, and the old man's head mushroom is about 3 / 5 - 4 / 5 of that of PC(18:2 / 18:2) in Tricholoma matsutake; The content of LPC(18:2) in Tricholoma matsutake, Russula brevipes, Flammulina velutipes, Agaricus blazei, and the old man's head mushroom is: Tricholoma matsutake > Agaricus blazei > Flammulina velutipes > the old man's head mushroom > Russula brevipes. The content of LPC(18:2) in Tricholoma matsutake is the highest, about 1.5 - 1.6 times that of LPC(18:2) in Agaricus blazei, about 2 times that of LPC(18:2) in Flammulina velutipes, about 2.2 - 2.25 times that of LPC(18:2) in the old man's head mushroom, and about 3.8 - 4 times that of LPC(18:2) in Russula brevipes; The content of LPE(18:2) in Tricholoma matsutake, Russula brevipes, Flammulina velutipes, Agaricus blazei, and the old man's head mushroom is: Tricholoma matsutake > Flammulina velutipes > Agaricus blazei > the old man's head mushroom > Russula brevipes. The content of LPE(18:2) in Tricholoma matsutake is the highest, about 2.6 - 2.7 times that of LPE(18:2) in Flammulina velutipes, about 3.4 - 3.5 times that of LPE(18:2) in Agaricus blazei, about 4.8 - 5 times that of LPE(18:2) in the old man's head mushroom, and about 37 times that of LPE(18:2) in Russula brevipes; The content of PA(18:2 / 18:2) in Russula brevipes is about 1.6 - 1.8 times that of PA(18:2 / 18:2) in Tricholoma matsutake. The content of PA(18:2 / 18:2) is comparable in Tricholoma matsutake, Agaricus blazei, and Flammulina velutipes, and the content of PA(18:2 / 18:2) in Tricholoma matsutake, Agaricus blazei, and Flammulina velutipes is: Tricholoma matsutake > Agaricus blazei > Flammulina velutipes. The content of PA(18:2 / 18:2) in Tricholoma matsutake is about 3.6 - 4 times that of PA(18:2 / 18:2) in the old man's head mushroom; The content of PA(16:0 / 18:2) in Tricholoma matsutake is relatively low, about 2 - 2.5 times, about 1 / 5 to 1 / 4 of the content of PA(16:0 / 18:2) in Russula alutacea, about 1 / 3 to 2 / 5 of the content of PA(16:0 / 18:2) in Agaricus blazei, about 3 / 4 to 4 / 5 of the content of PA(16:0 / 18:2) in Strobilomyces floccopus; the content of PE(18:2 / 18:2) in Tricholoma matsutake is the highest, equivalent to the content of PE(18:2 / 18:2) in Agaricus blazei, about 1.2 times the content of PE(18:2 / 18:2) in Agaricus blazei, about 2.7 to 3 times the content of PE(18:2 / 18:2) in Strobilomyces floccopus, about 3.9 to 4 times the content of PE(18:2 / 18:2) in Russula alutacea, about 12 to 12.5 times the content of PE(18:2 / 18:2) in Coprinus comatus; the content of DG(16:0 / 18:2) in Russula alutacea is much higher than that of the other four edible fungi. The contents of DG(16:0 / 18:2) in Russula alutacea, Strobilomyces floccopus, Agaricus blazei, Tricholoma matsutake, and Coprinus comatus are: Russula alutacea > Strobilomyces floccopus > Agaricus blazei > Tricholoma matsutake > Coprinus comatus. The content of DG(16:0 / 18:2) in Strobilomyces floccopus is about 9 / 50 to 1 / 5 times that of Russula alutacea, the content of DG(16:0 / 18:2) in Agaricus blazei is about 1 / 10 to 3 / 20 times that of Russula alutacea, the content of DG(16:0 / 18:2) in Tricholoma matsutake is about 1 / 11 to 1 / 10 times that of Russula alutacea, and the content of DG(16:0 / 18:2) in Coprinus comatus is about 1 / 25 that of Russula alutacea; the contents of TG(18:1 / 18:1 / 18:2) in Russula alutacea, Tricholoma matsutake, Strobilomyces floccopus, Agaricus blazei, and Coprinus comatus are: Strobilomyces floccopus > Tricholoma matsutake > Russula alutacea > Coprinus comatus > Agaricus blazei. The contents of TG(18:1 / 18:1 / 18:2) in Tricholoma matsutake and Russula alutacea are both about 2 to 2.2 times that in Agaricus blazei, the content of TG(18:1 / 18:1 / 18:2) in Coprinus comatus is about 1.5 to 1.6 times that in Agaricus blazei, and the content of TG(18:1 / 18:1 / 18:2) in Strobilomyces floccopus is about 3.6 to 3.65 times that in Tricholoma matsutake.

9. The identification method of Tricholoma matsutake and its similar edible fungi according to claim 1 or 7, characterized in that: DGTS(17:0 / 18:2) and PE(17:0 / 18:2) were only detected in Tricholoma matsutake and Strobilomyces floccopus. The content of DGTS(17:0 / 18:2) in Strobilomyces floccopus was about 5.9 - 6 times that in Tricholoma matsutake, and the content of PE(17:0 / 18:2) in Strobilomyces floccopus was about 1.8 - 2 times that in Tricholoma matsutake; PE(24:0 / 18:2) was only detected in Tricholoma matsutake and Agaricus blazei. The content of PE(24:0 / 18:2) in Agaricus blazei was about 5 - 5.5 times that in Tricholoma matsutake; Except for Lactarius volemus, PE(18:0 / 18:2) was detected in the other four edible fungi. The content of PE(18:0 / 18:2) in Tricholoma matsutake was the highest, about 2 - 2.2 times that in Strobilomyces floccopus, about 2.9 - 3 times that in Agaricus blazei, and about 4.8 - 5 times that in Russula brevipes; Except for Lactarius volemus, LPS(18:2) was detected in the other four edible fungi. The content of LPS(18:2) in Tricholoma matsutake was the highest, about 2 - 2.2 times that in Strobilomyces floccopus, about 4.5 times that in Agaricus blazei, and about 19 - 20 times that in Russula brevipes; The content of LPI(18:2) in Lactarius volemus was greater than that of PI(18:2) in the other four edible fungi. The content of LPI(18:2) in Russula brevipes, Tricholoma matsutake, Strobilomyces floccopus, Agaricus blazei, and Lactarius volemus was: Lactarius volemus > Strobilomyces floccopus > Tricholoma matsutake > Agaricus blazei > Russula brevipes. The content of LPI(18:2) in Lactarius volemus was about 1.5 - 1.55 times that in Strobilomyces floccopus, about 1.9 - 2 times that in Tricholoma matsutake, about 2.8 - 2.85 times that in Agaricus blazei, and about 8 times that in Russula brevipes.

10. A method for identifying matsutake and its similar edible fungi, characterized in that: Including: Step (1), extracting the lipids of Tricholoma matsutake and its similar edible fungi; Step (2), preparing the test sample solution, analyzing and identifying the lipid molecular structure by ultra - high performance liquid chromatography - high - resolution mass spectrometry, and constructing a standard curve through lipid standards of different lipid types to achieve quantitative analysis of lipid molecules in Tricholoma matsutake and its similar edible fungi; Based on the qualitative and quantitative results of lipid analysis, screening out differential lipid molecules through partial least squares analysis and p - value calculation; Step (3), extracting the lipids of Tricholoma matsutake and its similar edible fungi according to step (1), quantifying the screened differential lipid molecules according to step (2), and identifying the types of Tricholoma matsutake and its similar edible fungi based on the differential lipid molecules or the content of differential lipid molecules and differential lipid molecules.

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