Method for rapidly determining content of five artificial food spices in alcoholic drink

Abstract

The invention discloses a method for rapidly determining the content of five artificial food spices in alcoholic drink. The detection method is a gas chromatography-tandem mass spectrometry method. The method comprises the following steps: (1) weighing and transferring a beverage wine sample; (2) purifying and enriching the beverage wine sample by using a Shimazu SLE solid-phase column with diatomite as a filler; (3) carrying out constant volume filtration on the eluent; (4) setting instrument analysis conditions; (5) preparing a standard curve; and (6) calculating a result. The technology for detecting the five artificial food spices in the beverage wine is simple and efficient, the content of the five artificial food spices in the beverage wine can be measured only through one-time pretreatment and one-time on-machine testing, and the actual requirement for detection of the five common artificial food spices in the beverage wine can be better met.

Description

Technical Field

[0001] The invention relates to a method for rapidly determining the contents of five kinds of artificial food flavors in alcoholic beverages. Background Art

[0002] Artificial food flavors are flavor substances that are synthesized by chemical methods and are not found in natural products and are allowed to be eaten by humans. These flavors have significant flavoring and sweetening effects. For example, the sweetness of ethyl vanillin is 3 to 4 times that of vanillin, and the sweetness of ethyl maltol is 4 to 6 times that of maltol. Therefore, they are widely used in food, beverages, and alcoholic products. However, excessive addition of artificial food flavors will cause harm to the human body. Studies have shown that long-term consumption of foods with excessive ethyl maltol and ethyl vanillin will cause certain harm to human health, especially to human bones and liver. In severe cases, it may even cause symptoms such as difficulty breathing and vomiting. The American Flavor and Extract Manufacturers Association (FEMA) limits the use of ethyl maltol in wine to 18.6 mg / kg. The limit of ethyl vanillin in wine is 100 mg / kg. There has been some progress in the research on artificial food flavor detection technology. The country has also promulgated the detection method standards for ethyl maltol and ethyl vanillin in some foods, but there is no method standard for the detection of artificial food flavor content in beverage wine. At present, domestic scholars have conducted a lot of research on the aroma and flavor substances in beverage wine. The instrumental methods used mainly include: gas chromatography, gas chromatography-mass spectrometry, gas chromatography-tandem mass spectrometry, high performance liquid chromatography, high performance liquid chromatography-tandem mass spectrometry, Fourier transform infrared spectroscopy, etc. The detection of artificial food flavors in wine is reported only by high performance liquid chromatography and high performance liquid chromatography-tandem mass spectrometry, and the only artificial food flavor detected is ethyl maltol. Therefore, a method for quickly determining the content of 5 commonly used artificial food flavors in beverage wine is invented, which has important practical significance for the quality control of brewing enterprises and consumers' understanding of the quality of beverage wine products. Summary of the invention

[0003] The invention provides a method for rapidly determining the contents of five kinds of artificial food flavors in alcoholic beverages, which can solve the problem of the current single detection item of artificial food flavors in alcoholic beverages.

[0004] In order to solve the above problems, the present invention provides a method for quickly determining the content of five commonly used artificial food flavors in alcoholic beverages, wherein the five commonly used artificial food flavors are ethyl maltol, allyl cyclohexyl propionate, trans-myricetin, cis-myricetin, and ethyl vanillin. The specific steps are as follows: (1) Weighing and transfer of alcoholic beverages: Accurately weigh 2 g (accurate to 0.001 g) of the sample into a glass beaker and transfer it with a pipette to a Shimadzu solid phase SLE extraction column filled with diatomaceous earth placed on a vacuum extractor. Wash the beaker twice with 2 mL and 2 mL of distilled water, transfer the washing liquid to the extraction column, and let it stand for 10 min.

[0005] (2) Purification and enrichment of alcoholic beverage samples: After all samples are adsorbed on the filler, use a 50 mL stoppered glass graduated tube to receive the eluent, and use 25 mL of ethyl acetate to continuously elute the solid phase extraction column; (3) Volume adjustment and filtration of the eluent: After the eluent has completely flowed out, evacuate the solution for 1 min, add ethyl acetate to the volume of the receiving solution to 25 mL, shake well, filter through a 0.22 μm filter membrane, and then test on the machine; (4) Instrument conditions: Chromatographic conditions: Inlet temperature: 270 °C; Chromatographic column heating program: 80 °C for 1 min, 10 °C / min to 150 °C, 15 °C / min to 270 °C for 2 min. Carrier gas: Helium (purity not less than 99.999%); Flow rate: Constant flow mode, 1.0 mL / min; Injection volume 1 μL; Injection method: Splitless injection; Mass spectrometry conditions: interface temperature: 280°C; ion source: electron impact source (EI); ionization energy: 70 eV; ion source temperature: 230°C; quadrupole temperature: 150°C; solvent delay time: 4 min; monitoring mode: multiple reaction monitoring (MRM) mode, resolution of two quadrupole analyzers (MS1 and MS2): unit mass resolution; collision gas: high purity nitrogen; collision gas pressure: 200 kPa, collision gas flow rate: 0.55 mL / min. The mass spectrometry parameters of the target compound are shown in Table 1 Table 1 Mass spectrometry parameters of five artificial food flavors Note: The first product ion in Table 1 is the quantitative ion

[0006] 5) Preparation of standard curve: Weigh 10 mg (accurate to 0.1 mg) of standard substance, dissolve it in acetone and make up to 100 mL to prepare a single standard stock solution with a mass concentration of 100 μg / mL. 2.00 mL of myricetaldehyde standard stock solution, 1.00 mL of ethyl maltol standard stock solution, 1.00 mL of ethyl vanillin standard stock solution, and 1.00 mL of allyl cyclohexyl propionate standard stock solution were respectively transferred into a 100 mL volumetric flask, and the volume was fixed with acetone to prepare a mixed standard solution with a concentration of 1.00 μg / mL for trans-myricetaldehyde, cis-myricetaldehyde, ethyl maltol, ethyl vanillin, and allyl cyclohexyl propionate. The standard working solutions with concentrations of 5, 10, 50, 100, 500, and 1000 ng / mL were gradually diluted according to experimental needs. Samples were injected and analyzed according to step (4). The retention time and relative abundance ratio were used for qualitative analysis. The standard curve Y=AX+B was drawn based on the peak area of ​​the compound measured at each concentration and the corresponding mass concentration. (6) Calculation of results: Take the sample obtained in step (3) and perform analysis according to step (4). Measure the peak areas of the quantitative ions of the five artificial food flavoring compounds in the sample. Calculate the concentrations of the five artificial food flavoring compounds in the sample according to the standard curve prepared in (5). Further calculate the contents of the five artificial food flavoring compounds in the alcoholic beverage according to formula (1).

[0007] w = ---(1) Where: w──the content of five artificial food flavors in alcoholic beverages, in milligrams per kilogram (mg / kg); --The concentration of artificial food flavoring in alcoholic beverages calculated from the standard curve (μg / L); V – constant volume, in milliliters (mL); m – sample volume, in grams (g).

[0008] Compared with the prior art, the present invention has developed a method for rapidly determining the contents of five kinds of artificial food flavors in alcoholic beverages. The present invention has the following innovative features: The present invention relates to the accurate identification and quantification of 5 common artificial food flavors, involving more compounds and covering a wider range; The instrument used in the present invention is a gas chromatography-tandem triple quadrupole mass spectrometer, which has strong selectivity and anti-interference ability, high resolution and sensitivity for detecting compounds, low instrument cost and easier to promote; The present invention uses Shimadzu diatomaceous earth SLE solid phase extraction column for enrichment and purification, and after elution with ethyl acetate, innovatively adopts direct detection after passing through an organic phase filter membrane, without the need for concentration and other steps, thereby reducing the volatilization of the target compound and providing more accurate results. The present invention only requires one pre-treatment and one machine test to accurately analyze the contents of five kinds of artificial food flavors in alcoholic beverages, and can complete the detection of one sample within 15 minutes. BRIEF DESCRIPTION OF THE DRAWINGS

[0009] Figure 1 The following are the EI-MRM selected ion chromatograms of five artificial food flavorings (50 μg / L). DETAILED DESCRIPTION

[0010] The present invention is further described below by means of specific examples.

[0011] Example 1: Determination of the content of 5 artificial food flavors in prepared wine (1) Weighing and transfer of prepared liquor: Accurately weigh 2 g (accurate to 0.001 g) of the sample into a glass beaker and transfer it with a pipette to a solid phase extraction column filled with diatomaceous earth placed on a vacuum extractor. Wash the beaker twice with 2 mL and 2 mL of distilled water, transfer the washing liquid to the extraction column, and let it stand for 10 min.

[0012] (2) Purification and enrichment of blended liquor: After all samples are adsorbed on the filler, use a 50 mL stoppered glass graduated tube to receive the eluent, and use 25 mL of ethyl acetate to continuously elute the solid phase extraction column; (3) Volume adjustment and filtration of the eluent: After the eluent has completely flowed out, evacuate the solution for 1 min, add ethyl acetate to the volume of the receiving solution to 25 mL, shake well, filter through a 0.22 μm filter membrane, and then test on the machine; (4) Instrument conditions: Chromatographic conditions: Inlet temperature: 270 °C; Chromatographic column heating program: 80 °C for 1 min, 10 °C / min to 150 °C, 15 °C / min to 270 °C for 2 min. Carrier gas: Helium (purity not less than 99.999%); Flow rate: Constant flow mode, 1.0 mL / min; Injection volume 1 μL; Injection method: Splitless injection; Mass spectrometry conditions: interface temperature: 280°C; ion source: electron impact source (EI); ionization energy: 70 eV; ion source temperature: 230°C; quadrupole temperature: 150°C; solvent delay time: 4 min; monitoring mode: multiple reaction monitoring (MRM) mode, resolution of two quadrupole analyzers (MS1 and MS2): unit mass resolution; collision gas: high purity nitrogen; collision gas pressure: 200 kPa, collision gas flow rate: 0.55 mL / min. The mass spectrometry parameters of the target compound are shown in Table 1 5) Preparation of standard curve: Weigh 10 mg (accurate to 0.1 mg) of standard substance, dissolve it in acetone and make up to 100 mL to prepare a single standard stock solution with a mass concentration of 100 μg / mL. 2.00 mL of myricetaldehyde standard stock solution, 1.00 mL of ethyl maltol standard stock solution, 1.00 mL of ethyl vanillin standard stock solution, and 1.00 mL of allyl cyclohexyl propionate standard stock solution were respectively transferred into a 100 mL volumetric flask, and the solution was fixed to volume with acetone to prepare a mixed standard solution with a concentration of 1.00 μg / mL for trans-myricetaldehyde, cis-myricetaldehyde, ethyl maltol, ethyl vanillin, and allyl cyclohexyl propionate. The solution was diluted step by step according to experimental needs to obtain standard working solutions with concentrations of 5, 10, 50, 100, 500, and 1000 ng / mL. The samples were injected and analyzed according to step (4). The retention time and relative abundance ratio were used for qualitative analysis. The peak area of ​​the compound measured at each concentration and the corresponding mass concentration were used to draw a standard curve Y=AX+B (Y: peak area; X: mass concentration; A: slope; B: intercept); (6) Calculation of results: Take the sample obtained in step (3) and perform analysis according to step (4). Measure the peak areas of the quantitative ions of the five artificial food flavoring compounds in the blended alcoholic beverage. Calculate the concentrations of the five artificial food flavoring compounds in the blended alcoholic beverage according to the standard curve prepared in (5). Further, calculate the contents of the five artificial food flavoring compounds in the blended alcoholic beverage according to formula (1).

[0013] w = --(1) Where: w - the content of five artificial food flavors in the blended wine, in milligrams per kilogram (mg / kg); --The concentration of artificial food flavoring in the blended alcohol calculated from the standard curve (μg / L); V – constant volume, in milliliters (mL); m – sample volume, in grams (g).

[0014] The results showed that one artificial food flavoring, ethyl vanillin, was detected in the blended liquor at a content of 9.5 mg / kg, and the rest were not detected. See Table 2 for details.

[0015] Table 2 Content of artificial food flavors in blended alcoholic beverages

[0016] The above embodiments are only used to illustrate the technical solutions of the present invention rather than to limit it. Although the present invention has been described in detail with reference to the preferred embodiments, those skilled in the art should understand that the technical solutions of the present invention may be modified or replaced by equivalents without departing from the purpose and scope of the technical solutions of the present invention, which should all be included in the scope of the claims of the present invention.

Claims

1. A method for rapidly determining the content of five artificial food flavors in alcoholic beverages, characterized in that: The method for detecting the contents of five kinds of artificial food flavors in alcoholic beverages can complete the determination of the contents of five kinds of artificial food flavors in Table 1 with only one pretreatment and one machine operation. The specific steps are as follows: (1) Weighing and transfer of alcoholic beverages: Accurately weigh 2 g of the sample (accurate to 0.001 g) into a glass beaker and transfer it with a pipette to a Shimadzu SLE solid phase extraction column filled with diatomaceous earth placed on a vacuum extractor. Wash the beaker twice with 2 mL and 2 mL of distilled water respectively, transfer the washing liquid to the extraction column, and let it stand for 10 min. (2) Purification and enrichment of alcoholic beverage samples: After all samples are adsorbed on the filler, use a 50 mL stoppered glass graduated tube to receive the eluent, and use 25 mL of ethyl acetate to continuously elute the solid phase extraction column; (3) Volume adjustment and filtration of the eluent: After the eluent has completely flowed out, evacuate the solution for 1 min, add ethyl acetate to the volume of the receiving solution to 25 mL, shake well, filter through a 0.22 μm filter membrane, and then test on the machine; (4) Instrument conditions: Chromatographic conditions: Inlet temperature: 270 °C; Chromatographic column heating program: 80 °C for 1 min, 10 °C / min to 150 °C, 15 °C / min to 270 °C for 2 min. Carrier gas: Helium (purity not less than 99.999%); Flow rate: Constant flow mode, 1.0 mL / min; Injection volume 1 μL; Injection method: Splitless injection; Mass spectrometry conditions: interface temperature: 280°C; ion source: electron impact source (EI); ionization energy: 70 eV; ion source temperature: 230°C; quadrupole temperature: 150°C; solvent delay time: 4 min; monitoring mode: multiple reaction monitoring (MRM) mode, resolution of two quadrupole analyzers (MS1 and MS2): unit mass resolution; Collision gas: high purity nitrogen; collision gas pressure: 200 kPa, collision gas flow rate: 0.55 mL / min. The mass spectrometry parameters of the target compounds are shown in Table 1 (5) Preparation of standard curve: Weigh 10 mg (accurate to 0.1 mg) of standard substance, dissolve it in acetone and make up to 100 mL to prepare a single standard stock solution with a mass concentration of 100 μg / mL. 2.00 mL of myricetaldehyde standard stock solution, 1.00 mL of ethyl maltol standard stock solution, 1.00 mL of ethyl vanillin standard stock solution, and 1.00 mL of allyl cyclohexyl propionate standard stock solution were respectively transferred into a 100 mL volumetric flask, and the volume was fixed with acetone to prepare a mixed standard solution with a concentration of 1.00 μg / mL for trans-myricetaldehyde, cis-myricetaldehyde, ethyl maltol, ethyl vanillin, and allyl cyclohexyl propionate. The standard working solutions with concentrations of 5, 10, 50, 100, 500, and 1000 ng / mL were gradually diluted according to experimental needs. Samples were injected and analyzed according to step (4). The retention time and relative abundance ratio were used for qualitative analysis. The standard curve Y=AX+B was drawn based on the peak area of ​​the compound measured at each concentration and the corresponding mass concentration. (6) Calculation of results: Take the sample obtained in step (3) and analyze it according to step (4). Measure the peak area of ​​the quantitative ions of the five artificial food flavoring compounds in the sample. Calculate the concentration of the five artificial food flavoring compounds in the sample according to the standard curve prepared in (5). Further calculate the content of the five artificial food flavoring compounds in the beverage according to formula (1): w = （1） Where: w──the content of five artificial food flavors in alcoholic beverages, in milligrams per kilogram (mg / kg); --The concentration of artificial food flavoring in alcoholic beverages calculated from the standard curve (μg / L); V – constant volume, in milliliters; m – sample volume, in grams.

2. The method for determining the contents of five artificial food flavors in alcoholic beverages according to claim 1, characterized in that: The solid phase extraction column described in (1) is a Shimadzu SLE solid phase extraction column (10 mL, 5 g) filled with diatomaceous earth. After the sample is transferred to the solid phase extraction column, the beaker is washed twice with 2 mL of distilled water, and the washing liquid is transferred to the extraction column. The column is allowed to stand for 10 minutes until the sample solution is completely adsorbed.

3. The method for determining the contents of five artificial food flavors in alcoholic beverages according to claim 1, characterized in that: The elution solvent described in (2) is ethyl acetate, and the volume of the elution solvent is 25 mL. During the elution process, it is necessary to ensure that the liquid surface of the solid phase column does not dry up.

4. The method for determining the contents of five artificial food flavors in alcoholic beverages according to claim 1, characterized in that: After elution with ethyl acetate solvent as described in (3), vacuum is applied for 1 min. The receiving solution is made up to volume with ethyl acetate and shaken well, and then filtered with a 0.22 μm organic phase filter membrane.

5. The method for determining the contents of five artificial food flavors in alcoholic beverages according to claim 1, characterized in that: The chromatographic column described in (4) is a ZB-1701 quartz capillary column (30 m × 0.25 mm × 0.25 μm) (Phenomenex, USA), the ion source is an electron impact source (EI), and the collision gas is high-purity nitrogen.

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