Method for detecting sweet-scented osmanthus aroma components in sweet-scented osmanthus wine

Abstract

The invention discloses a method for detecting sweet-scented osmanthus aroma components in sweet-scented osmanthus wine, which comprises the following steps: diluting a wine sample, adding salt until the wine sample is saturated, and then carrying out liquid-liquid micro-extraction by using a mixed solvent of absolute ether and ethyl acetate in a specific ratio, the volume fraction of the ethyl acetate being 3-5%; after oscillation, ultrasonic-assisted extraction and centrifugal separation, an organic phase is taken for dehydration, a gas chromatograph-mass spectrometer is used for analysis, and an internal standard method is adopted for quantification. By optimizing the polarity of the solvent, various aroma components (such as leaf alcohol, linalool oxide, linalool, ionone, tea aroma spirane and the like) from strong polarity to weak polarity can be effectively covered, and balanced and efficient extraction under a single extraction step is realized. The method is easy and convenient to operate, high in recovery rate, good in reproducibility and suitable for quality evaluation and production process control of the osmanthus fragrans wine.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of food flavor detection, and in particular to a method for detecting a plurality of osmanthus flower aroma components in osmanthus flower wine by using gas chromatography-mass spectrometry (GC-MS). BACKGROUND

[0002] Osmanthus flower wine is a flavor wine made by adding osmanthus flowers to different wine bases through processes such as steeping, re-distilling, or mixed distilling. Among them, osmanthus-flavored liquor made by using rice-flavored liquor as the base liquor through specific processes is an important representative category. Regardless of the specific base liquor and process, the typical osmanthus flower aroma is an important indicator for evaluating the quality of the product. This aroma is derived from a series of volatile compounds, mainly including terpene alcohols (such as linalool, geraniol), terpene oxides (such as linalool oxide), ketones (such as alpha-ionone, beta-ionone), lactones and esters (such as gamma-decalactone, ethyl phenylacetate), and tea squalane and other terpene derivatives. Therefore, it is necessary to develop a pretreatment method that can better adapt to the aroma components with a wide range of polarities in osmanthus flower wine to achieve accurate quantitative analysis.

[0003] At present, gas chromatography-mass spectrometry is a commonly used means for analyzing such volatile components, and its accuracy depends largely on the sample pretreatment effect. Liquid-liquid extraction is a commonly used pretreatment method, but when applied to the above-mentioned aroma components, it faces a practical problem: the physical and chemical properties of these components differ greatly, and their octanol-water partition coefficients cover a wide range (for example, the LogP value of leaf alcohol is about 1.3, while the LogP value of tea squalane is significantly higher than 4.0). If a single polarity traditional extraction solvent is used, it is often difficult to achieve balanced and efficient extraction of components with a wide range of polarities, which may result in suboptimal recovery of some components or poor extraction selectivity, thereby causing deviations in the quantitative results. Therefore, a pretreatment method that can better adapt to components with a wide range of polarities is needed to support accurate quantitative analysis. SUMMARY

[0004] To solve the above technical problems, the present application provides a pretreatment method and a GC-MS detection method which are simple to operate and can achieve balanced and efficient extraction of osmanthus flower aroma components with a wide range of polarities in a single extraction step. This method is aimed at the wide range of polarities of the aroma components in such wine samples, and is designed to optimize the extraction solvent system in the pretreatment process to improve the problem of uneven extraction efficiency of different polarity components when using a single solvent, so as to better cover the main target components in one treatment and support accurate quantitative analysis.

[0005] To achieve the above-mentioned purposes, the present application adopts the following technical solutions: A method for detecting osmanthus flower aroma components in osmanthus flower wine, comprising the following steps: S1. Sample preparation and internal standard addition: Take the wine sample to be tested, add the mixed internal standard solution, and dilute with ultrapure water to the specified ethanol volume fraction to reduce matrix interference.

[0006] S2. Salt addition and solvent addition: Add sodium chloride to the solution obtained in step S1 to saturation, and then add the extraction solvent.

[0007] In this step, sodium chloride is added to the obtained solution to saturation, and the solubility of the target aroma components in the aqueous phase is reduced by using salting-out effect, promoting their partitioning into the organic extraction solvent.

[0008] S3. Extraction and separation: Shake the solution system obtained in step S2, perform ultrasonic-assisted extraction under low temperature conditions, then centrifuge, and after the two phases are fully separated, take the upper organic phase to obtain the test solution.

[0009] S4. Gas chromatography-mass spectrometry analysis and quantification: The test solution obtained in step S3 is analyzed by GC-MS, and the internal standard standard curve method is used to calculate the content of the target component.

[0010] Preferably, in step S1, the ultrapure water is diluted to an ethanol volume fraction of 7%-15%, preferably 9% to 12%.

[0011] Preferably, the mixed internal standard solution in step S1 contains anisyl acetone, alpha-terpineol-d3, L-menthol, and octanoic acid ethyl ester-d15.

[0012] Preferably, in step S2, the extraction solvent is a mixture of anhydrous diethyl ether and ethyl acetate, wherein the volume fraction of ethyl acetate is 1% to 10%, preferably 3% to 5%.

[0013] Preferably, before step S1, a standard curve is prepared: a series of gradient concentration standard solutions are prepared and internal standards are added; the internal standard standard curve method is prepared by analyzing the series of standard solutions and the wine sample to be tested simultaneously after the complete pretreatment of steps S2 to S3, and then analyzed by step S4. More preferably, the series of standard solutions are prepared using ethanol aqueous solution with similar ethanol concentration as the sample pretreatment to simulate the matrix and reduce the quantitative error.

[0014] Preferably, the GC-MS analysis conditions in step S4 include: the chromatographic column is a nitroterephthalic acid modified polyethylene glycol capillary column; the temperature programming is 35-45 ℃ for 1-3 min, then 8-12 ℃ / min to 85-95 ℃ for 2-4 min, then 2-4 ℃ / min to 200-220 ℃, finally 8-12 ℃ / min to 235-245 ℃ for 4-6 min; the mass spectrometer uses an electron impact source.

[0015] Preferably, the internal standard standard curve method in step S4 adopts a matrix-matched standard curve for quantification, which is prepared by formulating a series of gradient concentrations of standard solutions and adding an internal standard, synchronously processing the series of standard solutions using the same pretreatment method as the wine sample to be tested, and then plotting a standard curve by GC-MS analysis.

[0016] Preferably, in step S4, the internal standard standard curve method is used to calculate the content of the target component, and the calculation formula is:

[0017] In the formula, X i is the content of the ith target, with units of μg / L; X f is the known content of the corresponding internal standard, with units of μg / L; A i is the chromatographic peak area of the target; A f is the chromatographic peak area of the internal standard; b 、 k is the intercept and slope of the standard curve; K is the dilution factor.

[0018] Preferably, the osmanthus fragrance components include at least ten of leaf alcohol, furan oxidized linalool (isomer mixture), tea-scented spiro compound (isomer mixture), linalool, alpha-terpineol, pyran oxidized linalool (isomer mixture), (+)-beta-citronellol, ethyl benzoate, geraniol, dihydro-beta-ionone, geranial, alpha-ionone, alpha-ionol, beta-ionone, nerolidol (isomer mixture), and gamma-decalactone.

[0019] Compared with the prior art, the present application has the following beneficial effects: (1) The method can extract fragrance components with a wide polarity range: By optimizing the solvent polarity (such as 3%-5%), the extraction of multiple fragrance components with different polarities can be considered at the same time. A small amount of ethyl acetate helps to improve the extraction of strong polarity components (such as leaf alcohol and oxidized linalool), and the main part of anhydrous diethyl ether maintains good extraction ability for medium and weak polarity components (such as linalool and ionone) and strong hydrophobic components (such as tea-scented spiro compound and nerolidol). This design enables the method to cover a wide range of compounds with different polarities.

[0020] (2) The quantitative result is accurate: the method requires the complete pretreatment of the series of standard solutions and the sample to be tested synchronously, and the standard curve is drawn by using the matrix matched standard solution. This method helps to reduce the quantitative deviation caused by the complexity of the sample matrix and the pretreatment process, thereby supporting to obtain more reliable analysis results.

[0021] (3) The method is stable and practical: the method steps are clear and the operation is relatively simple. Through sample dilution, salt addition, oscillation and low temperature ultrasonic assisted extraction, etc., the recovery rate and operation reproducibility of the method are improved. The established method is suitable for quality analysis and product evaluation in the production process of osmanthus wine. BRIEF DESCRIPTION OF DRAWINGS

[0022] In order to more clearly illustrate the technical solutions of the embodiments of the present application or the prior art, the drawings needed to be used in the embodiments or the prior art description will be briefly introduced below. Obviously, the drawings in the following description are only some embodiments of the present application, and other drawings can also be obtained by those skilled in the art without creative labor.

[0023] Figure 1 The chromatogram of the mixed standard solution of 16 aroma components and the internal standard in example 1; Figure 2 The standard curve diagram of leaf alcohol in example 1; Figure 3 The standard curve diagram of furan class of oxidized linalool in example 1; Figure 4 The standard curve diagram of tea aroma spirans in example 1; Figure 5 The standard curve diagram of linalool in example 1; Figure 6 The standard curve diagram of alpha-terpineol in example 1; Figure 7 The standard curve diagram of pyran class of oxidized linalool in example 1; Figure 8 The standard curve diagram of (+)-beta-citronellol in example 1; Figure 9 The standard curve diagram of ethyl phenylacetate in example 1; Figure 10 The standard curve diagram of nerol in example 1; Figure 11 The standard curve diagram of dihydro-beta-ionone in example 1; Figure 12 The standard curve diagram of geraniol in example 1; Figure 13 The standard curve diagram of alpha-ionone in example 1; Figure 14 Standard curve plot for a-lonone in Example 1; Figure 15 Standard curve plot for β-damascenone in Example 1; Figure 16 Standard curve plot for nerolidol in Example 1; Figure 17 Standard curve plot for γ-decalactone in Example 1; Figure 18 Chromatogram of Sample 1 of Example 1; Figure 19 Chromatogram of recovery experiment of Sample 1 of Example 1. DETAILED DESCRIPTION

[0024] The technical solutions in the embodiments of the present application will be clearly and completely described below in conjunction with the embodiments of the present application. Obviously, the described embodiments are only part of the embodiments of the present application, rather than all the embodiments of the present application. Based on the embodiments of the present application, all other embodiments obtained by those of ordinary skill in the art without creative work fall within the scope of protection of the present application.

[0025] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the embodiments of the present application belong. If the definitions stated in this section are contrary to or otherwise inconsistent with the definitions stated in the patents, patent applications, published patent applications, and other publications that are cited herein by reference, the definitions stated in this section take precedence over the definitions stated in the cited references.

[0026] The methods used in the following examples are conventional unless otherwise stated. The materials, reagents and instruments used are conventional in the art unless otherwise stated, and are available to those skilled in the art through commercial channels.

[0027] When a quantity, concentration, or other value or parameter is expressed as a range, a preferred range, or a range defined by a series of upper and lower preferred values, this should be understood as specifically disclosing all ranges formed by any pair of any upper or preferred value with any lower or preferred value, regardless of whether the range is disclosed individually. For example, when the range “1 to 5” is disclosed, the described range should be interpreted as including ranges “1 to 4”, “1 to 3”, “1 to 2”, “1 to 2 and 4 to 5”, “1 to 3 and 5”, etc. When numerical ranges are described herein, unless otherwise stated, the range is intended to include its endpoints and all integers and fractions within that range. In this specification and claims, range definitions may be combined and / or interchanged, unless otherwise stated, these ranges include all subranges contained therein.

[0028] The present invention will be further described below with reference to examples and comparative examples. The osmanthus aroma components to be detected in the present invention have a wide distribution of lipophilicity (expressed as octanol-water partition coefficient LogP). Specifically, the lipophilicity of these components varies significantly: (1) Strongly polar components: such as linalool, linalool oxide, etc., have low LogP values ​​(<2.0) and strong hydrophilicity; (2) Strongly or moderately polar components: such as linalool, geraniol, ionone, etc.; (3) Weakly polar components: such as tea spiroane, nerolidol, etc., have high LogP values ​​(>4) and significant hydrophobicity. The present invention is illustrated with a typical osmanthus wine, but the method is also applicable to osmanthus wine made by other processes.

[0029] Example 1 This embodiment provides a quantitative detection method for 16 osmanthus aroma components in osmanthus-flavored baijiu, as detailed below: 1. Reagents Anhydrous ethanol, anhydrous diethyl ether, and ethyl acetate were of chromatographic grade; anhydrous Na2SO4 and NaCl were of analytical grade; and water was ultrapure water.

[0030] 2. Samples to be tested and standards Sample 1 to be tested: provided by Tianlongquan Winery; Standards: Leaf alcohol, furan-derived linalool oxide (mixture of isomers), tea spiroane (mixture of isomers), linalool, α-terpineol, pyran-derived linalool oxide (mixture of isomers), (+)-β-citronellol, ethyl phenylacetone, nerol, dihydro-β-ionone, geraniol, α-ionone, α-ionol, β-ionone, nerolidol (mixture of isomers), γ-decanolide, anisylacetone, α-terpineol-d3, L-menthol and ethyl caprylate-d 15The purity of α-ionone and α-ionol is greater than 90%, and the purity of the remaining standard substances is greater than 95%, all purchased from Sigma-Aldrich Company, USA.

[0031] 3. Preparation of standard solution 3.1 Preparation of mixed standard stock solution The above 16 kinds of aroma substance standard samples were precisely weighed into 10 mL volumetric flasks, anhydrous ethanol was added to the calibration mark to obtain a single standard stock solution, and then an appropriate amount of single standard stock solution was removed into 10 mL volumetric flasks, anhydrous ethanol was added to the calibration mark, and it was stored at 4°C in the dark. The specific concentration is shown in Table 1.

[0032] Table 1 Concentrations of 16 kinds of aroma component single standard solution and mixed standard solution

[0033] 3.2 Internal standard solution Anisyl acetone, α-terpineol-d3, L-menthol and ethyl octanoate-d 15 were precisely weighed into 10 mL volumetric flasks, dissolved and diluted to the calibration mark with anhydrous ethanol to obtain a mixed internal standard solution, which was stored at 4°C in the dark. The concentrations of anisyl acetone, α-terpineol-d3, L-menthol and ethyl octanoate-d 15 were 450, 400, 400 and 500 mg / L, respectively.

[0034] 4. Sample pretreatment Take 4 mL of wine sample (55%vol) in a 50 mL centrifuge tube, add 10 μL of internal standard, add 18 mL of ultrapure water (ethanol volume fraction is 10%), add 7.2 g of sodium chloride to saturate the solution, then add 2 mL of a mixed solvent of anhydrous ether and ethyl acetate (the volume fraction of ethyl acetate is 5%), shake for 5 min, ultrasonic in cold water for 10 min, centrifuge at 8000 rpm for 5 min, stand at 4°C for more than 12 hours (to promote the full separation of the two phases), take the upper organic phase, dehydrate with anhydrous Na2SO4, and the obtained sample solution is detected by gas chromatography-mass spectrometry.

[0035] 5. Sample detection 5.1 Instrument conditions Instrument: Agilent gas chromatography-mass spectrometry (8890B-5977BMSD).

[0036] Chromatographic conditions: A nitroterephthalic acid modified polyethylene glycol capillary chromatographic column (DB-FFAP: 60 m x 0.25 mm x 0.25 μm) was used, the injection port temperature was 250 °C, the carrier gas was high-purity helium (purity ≥ 99.999%), non-split constant flow mode injection, the flow rate was 1 mL / min, the injection amount was 1 μL, and the temperature program was as follows: the initial temperature was 40 °C, maintained for 2 min, increased to 90 °C at a rate of 10 °C / min, maintained for 3 min, increased to 210 °C at a rate of 3 °C / min, maintained for 1 min, increased to 240 °C at a rate of 10 °C / min, and maintained for 5 min.

[0037] Mass spectrometric conditions: The ionization mode was electron impact source (EI) with an energy of 70 eV, the ion source temperature was 230 °C, the quadrupole rod temperature was 150 °C, the auxiliary channel heating temperature was 280 °C, and the mass spectrometric scanning mass range was 50-350 amu. The mass spectrometric analysis used NIST20 spectral library.

[0038] Detection mode: selected ion scanning collection; the monitoring ions are shown in Table 2. Scanning mode: first, full scan mode (SCAN) was used for qualitative analysis, the instrument parameters were optimized to determine the retention time and characteristic ions of each effective component and the internal standard, then the compounds were grouped, and selected ion mode (SIM) scanning was used, as shown in Table 2.

[0039] Table 2 Information summary table of retention time and internal standard quantification of 16 aroma components

[0040] 5.2 Preparation of standard curve According to “Table 1” in “3.1”, an appropriate amount of 16 kinds of aroma component mixed standard solution stock solution was taken in a 10 mL volumetric flask, and the same alcohol content (55% vol) ethanol solution as the sample was used to make up the sample 1, and then 2.5 times of the sample 1 was diluted to obtain a series of mixed standard solutions. The pretreatment was carried out according to the method in “4”, and then the gas chromatography-mass spectrometry was used for detection. The peak area ratio of the test substance to the corresponding internal standard was taken as the ordinate, and the concentration ratio was taken as the abscissa, linear regression was carried out, and the standard curve was drawn. For furan and pyran linalool isomer mixture, tea aroma spiranes isomer mixture and nerolidol isomer mixture, the peak areas of the cis and trans isomers were added together, and the total concentration of the mixture was used for the preparation of the standard curve and sample calculation. The standard curve parameters of the 16 kinds of aroma components are shown in Table 3, and the standard curves are shown in Figures 2-17 , Figure 1 The chromatographic peaks 1-24 in Table 3 are leaf alcohol, ethyl octanoate d 15Furanoid linalool oxide isomer 1, Furanoid linalool oxide isomer 2, Tea squalene isomer 1, Tea squalene isomer 2, Linalool, L-Menthol, Alpha-Terpineol-d3, Alpha-Terpineol, Pyranoid linalool oxide isomer 1, Pyranoid linalool oxide isomer 1, (+)-Beta-Myrcenol, Ethyl benzoate, Nerol, Dihydro-beta-ionone, Geraniol, Alpha-ionone, Alpha-ionol, Beta-ionone, Nerolidol isomer 1, Nerolidol isomer 2, Gamma-decalactone, Anisyl acetone. The limit of detection (LOD) is the sample concentration corresponding to a signal-to-baseline noise (S / N) of 3:1; the limit of quantitation (LOQ) is the sample concentration corresponding to a signal-to-baseline noise (S / N) of 10:1.

[0041] Table 3 Summary of standard curve parameters for 16 aroma components

[0042] 5.3 Sample determination and calculation The sample solution and the standard solution were determined under the same detection conditions, and the peak area A i of the 16 aroma components and the peak area A f of the internal standard were substituted into the standard curve to calculate the corresponding mass concentration X i of the sample. The chromatogram of the sample to be tested is shown in Figure 18 .

[0043] The content of the 16 aroma substances in the sample to be tested is calculated according to the following formula

[0044] In the formula, X i is the content of the i-th target substance, in units of micrograms per liter (μg / L); X f is the content of the corresponding internal standard (μg / L); A i and A f are their chromatographic peak areas, respectively; b and k are the intercept and slope of the standard curve; and K is the dilution factor. The calculation results are expressed as the arithmetic mean of two independent determination results obtained under repeatability conditions, and are retained to two decimal places.

[0045] 6. Repeatability experiment Six samples to be tested were taken, labeled 1-6, and the samples were processed according to the methods of "4. Sample pretreatment" and "5. Sample detection", the content of 16 aroma components was determined, and the repeatability of the determination results of each aroma component was calculated. The RSD values were all less than 10%, indicating that the method had good repeatability, and the specific results are shown in Table 4.

[0046] Table 4 Repeatability experiment results of 16 aroma components in wine samples

[0047] 7. Accuracy experiment Take 6 samples of the wine to be tested, divide them into A, B, C, a total of 3 groups, 2 samples in each group, and mark them as A group (Z1-1 and Z1-2), B group (Z2-1 and Z2-2), and C group (Z3-1 and Z3-2). Take the 16 compound mixed standard stock solution in “3.1”, and add 10 μL, 20 μL and 30 μL to the A group, B group and C group respectively. Process the samples according to the methods of “4. Sample pretreatment” and “5. Sample detection”, and determine the content of the target substances, as shown in Table 5, calculate the content and recovery rate of each compound, the recovery rate of 16 target substances is between 85% and 110%, and the RSD value is less than 10%, indicating that the accuracy of the method is good, and the specific calculation results are shown in Table 5. Figure 19

[0048] Table 5: Recovery rate experiment results of 16 aroma components in wine samples

[0049] Example 2 The present embodiment provides a method for detecting the concentration of 16 aroma components in Gui Xiang type Baijiu, which is as follows: 1. Reagents: The same as in Example 1.

[0050] 2. Test sample and standard: Test sample 2: provided by Tianlongquan Liquor Co., Ltd., alcohol content 55%vol, Standard: The same as in Example 1.

[0051] 3. Preparation of standard solution: The same as in Example 1.

[0052] 4. Sample pretreatment: Except that the extraction solvent is changed to a mixture of anhydrous ether and ethyl acetate (the volume fraction of ethyl acetate is 3%), the rest of the steps are the same as in Example 1.

[0053] 5. Sample detection: The same as in Example 1.

[0054] The detection results of 16 aroma components in the test sample 2 of the present embodiment are shown in Table 6.

[0055] Table 6: Detection results of 16 aroma components in wine samples

[0056] Comparative Example 1: Using pure anhydrous ether as extraction solvent 1. Reagents: The same as in Example 1.

[0057] 2. Test sample and standard: Test sample 1: provided by Tianlongquan Liquor Co., Ltd., alcohol content 55%vol.​

[0058] Standard product: Same as in Example 1.

[0059] 3. Preparation of standard solutions: Same as in Example 1.

[0060] 4. Sample pretreatment: Except for changing the extraction solvent to pure anhydrous diethyl ether, the other steps are the same as in Example 1.

[0061] 5. Sample testing: Same as in Example 1.

[0062] 6. Accuracy test: Same as in Example 1.

[0063] The test results and recovery rate results of sample 1 in this comparative example are shown in Table 7.

[0064] Table 7. Results of the spiked recovery of 16 aroma components in wine samples (anhydrous diethyl ether)

[0065] Results: The average recovery rate of the highly polar component leaf alcohol was only 75.62%, while the recovery rate of the weakly polar component tea spirolane was 95.21%. This indicates that pure diethyl ether has insufficient extraction capacity for highly polar components.

[0066] Comparative Example 2: Using a mixed solvent containing 20% ​​ethyl acetate 1. Reagents: Same as in Example 1.

[0067] 2. Samples to be tested and standards: Sample 1 to be tested: provided by Tianlongquan Winery, alcohol content 55% vol.

[0068] Standard product: Same as in Example 1.

[0069] 3. Preparation of standard solutions: Same as in Example 1.

[0070] 4. Sample pretreatment: Except for changing the extraction solvent to a mixture of anhydrous diethyl ether and ethyl acetate (ethyl acetate volume fraction of 20%), the other steps are the same as in Example 1.

[0071] 5. Sample testing: Same as in Example 1.

[0072] 6. Accuracy test: Same as in Example 1.

[0073] The test results and recovery rate of sample 1 in this comparative example are shown in Table 8.

[0074] Table 8. Results of the spiked recovery of 16 aroma components in wine samples (20% ethyl acetate)

[0075] Results: Spiked recovery experiments showed that the recovery rate of leaf alcohol was 94.83%, the recovery rate of linalool was 92.15%, while the recovery rate of tea spiroane decreased to 80.21%. These results indicate that while a high ethyl acetate ratio can effectively extract strongly polar components, it leads to a decrease in the recovery rate of weakly polar components, also causing uneven extraction.

[0076] Comparison of results between the examples and the comparative examples Table 9 Comparison of extraction efficiencies (recoveries) of aroma components with a wide polarity range using different solvent systems.

[0077] In summary, the data in the table above shows that the solvent systems of Comparative Example 1 (pure anhydrous diethyl ether) and Comparative Example 2 (high proportion of ethyl acetate) both exhibit a certain degree of bias in their extraction efficiency for different polar components. In contrast, Example 1, using a formulation containing 3%-5% ethyl acetate, achieves a better balance between the extraction efficiency of strongly polar and weakly polar components, thus enabling a more comprehensive and balanced extraction of aroma components with a wide range of polarities in a single treatment. This indicates that this specific solvent formulation is of significant value in improving extraction results and solving the technical problem of this application.

[0078] The above embodiments are merely illustrative examples of preferred embodiments of the present invention and do not encompass all possible implementations. Any modifications and refinements made by those skilled in the art without departing from the spirit and scope of the present invention are considered to fall within the protection scope of the claims.

Claims

1. A method for detecting osmanthus aroma components in osmanthus wine, characterized in that, Includes the following steps: S1. Sample preparation and internal standard addition: Take the wine sample to be tested, add the mixed internal standard solution, and dilute with ultrapure water to the specified ethanol volume fraction; S2. Adding salt and solvent: Add sodium chloride to the solution obtained in step S1 until saturated, and then add the extraction solvent; S3. Extraction and separation: The solution system obtained in step S2 is shaken and ultrasonic-assisted extraction is performed at low temperature. Then, centrifugation is performed. After the two phases are fully separated, the upper organic phase is taken and dehydrated to obtain the test solution. S4. Gas Chromatography-Mass Spectrometry Analysis and Quantification: The test solution obtained in step S3 was analyzed by GC-MS, and the content of the target component was calculated using the internal standard curve method.

2. The method according to claim 1, characterized in that, In step S1, the ethanol is diluted with ultrapure water to a volume fraction of 7%-15%, preferably 9% to 12%.

3. The method according to claim 1, characterized in that, The mixed internal standard solution in step S1 contains anisylacetone, α-terpineol-d3, L-menthol and ethyl octanoate-d15.

4. The method according to claim 1, characterized in that, The extraction solvent in step S2 is a mixture of anhydrous diethyl ether and ethyl acetate, wherein the volume fraction of ethyl acetate is 1% to 10%, preferably 3% to 5%.

5. The method according to claim 1, characterized in that, Before step S1, the preparation of a standard curve is also included: a series of standard solutions with gradient concentrations are prepared and an internal standard is added; the internal standard curve method is to draw the curve by simultaneously performing the series of standard solutions and the wine sample to be tested on the complete pretreatment in steps S2 to S3, and then analyzing and plotting it in step S4; more preferably, the series of standard solutions are prepared using an ethanol-water solution with a concentration similar to that of the sample during pretreatment, in order to simulate the matrix and reduce quantitative error.

6. The method according to claim 1, characterized in that, The GC-MS analysis conditions in step S4 include: the chromatographic column is a nitroterephthalic acid modified polyethylene glycol capillary column; the temperature program is as follows: initial temperature 35-45 ℃ held for 1-3 min, increased to 85-95 ℃ at 8-12 ℃ / min and held for 2-4 min, then increased to 200-220 ℃ at 2-4 ℃ / min, and finally increased to 235-245 ℃ at 8-12 ℃ / min and held for 4-6 min; the mass spectrometer uses an electron impact source.

7. The method according to claim 1, characterized in that, In step S4, the internal standard curve method uses a matrix-matched standard curve for quantification. The matrix-matched standard curve is prepared by preparing a series of standard solutions with gradient concentrations and adding an internal standard. After simultaneously processing the series of standard solutions using the same pretreatment method as the wine sample to be tested, the curve is plotted by GC-MS analysis.

8. The method according to claim 1, characterized in that, In step S4, the content of the target component is calculated using the internal standard curve method, and the calculation formula is as follows: In the formula, X i The concentration of the i-th target compound is expressed in μg / L. X f The concentration of the corresponding internal standard is known, in μg / L; A i The peak area of ​​the target analyte; A f The chromatographic peak area of ​​the internal standard; b , k The intercept and slope of the standard curve; K This represents the dilution factor.

9. The method according to claim 1, characterized in that, The osmanthus fragrance components include at least ten of the following: leaf alcohol, a mixture of isomers of furan-based linalool oxide, a mixture of isomers of tea spiroane, linalool, α-terpineol, a mixture of isomers of pyran-based linalool oxide, (+)-β-citronellol, ethyl phenylacetate, nerol, dihydro-β-ionone, geraniol, α-ionone, α-ionol, β-ionone, a mixture of isomers of nerolidol, and γ-decanolide.

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