Detection method for rapidly determining 17 flavor enhancers in tea leaves

Abstract

The invention discloses a detection method for rapidly determining 17 flavor enhancers in tea leaves. The method comprises the following steps: (1) preparing a tea leaf sample; (2) extracting a tea sample; (3) purifying the tea sample; (4) setting instrument analysis conditions; (5) preparing a standard curve; and (6) calculating a result. The 17 flavor enhancers involved in the method are more in variety and more complete, the method is simple and efficient, the content of the 17 flavor enhancers in a tea sample can be determined only through one-time pretreatment and one-time machine testing, and the actual requirement for detection of the 17 flavor enhancers in tea can be better met.

Description

Technical Field

[0001] The invention relates to a detection method for rapidly determining 17 flavor enhancers in tea. Background Art

[0002] Tea aroma is one of the important qualities of tea. Tea of ​​different varieties and origins has its own unique aroma. Appendix B of my country's national food safety standard GB 2760-2014 "National Food Safety Standard for the Use of Food Additives" clearly lists tea as a food that cannot be added with food flavors and spices. However, there is currently a lack of detection methods for banned spices in tea. Therefore, the invention of a method that can simultaneously and quickly determine 17 flavor enhancers in tea can not only provide technical support for safety supervision in the tea field, but also has great significance for ensuring the quality and safety of tea products and promoting the healthy development of the tea industry. Summary of the invention

[0003] The task of the present invention is to provide a method for rapidly determining 17 flavor enhancers in tea, which can solve the problem that when detecting the content of flavor enhancers in multiple teas, multiple methods need to be used for simultaneous determination, which is time-consuming and requires the consumption of a large amount of organic solvents.

[0004] In order to solve the above problems, the present invention provides a method for rapidly and simultaneously determining 17 flavor enhancers in tea. The names of the 17 flavor enhancers are as listed in Table 1. The specific steps are as follows: (1) Preparation of tea samples: Take an appropriate amount of representative sample, grind it with a grinder, pass it through a 40-mesh sieve, put it into a clean container, seal it and label it; (2) Extraction of tea samples: Weigh 2 g of the sample in step (1) into a 50 mL centrifuge tube, accurately add 10 mL of acetonitrile, vortex for 1 min, and then place in an ultrasonic extractor for extraction for 30 min; (3) Purification of tea samples: The sample solution obtained in step (2) was activated (with 6.0 mL acetonitrile and 6 mL ultrapure water, respectively) and purified by PRiME HLB solid phase extraction column (6 mL, 500 mg). The eluate from the sample passing through the PRiME HLB solid phase extraction column was directly collected and filtered through a 0.22 μm PTFE filter membrane for testing; (4) Setting of instrument analysis conditions: The chromatographic column was a WATERS ACQUITY UPLC HSS T3 C18 column (1.7 µm, 2.1 x 150 mm); column temperature: 40 °C; flow rate: 0.3 mL / min; injection volume: 2 µL; mobile phase A was acetonitrile, and mobile phase B was 0.1% formic acid in water; gradient elution program: 0-4.5 min, 25% B-90% B; 4.5-6.5 min, 90% B; 6.5-7 min, 90% B-25% B; 7-9 min, 25% B; ion source was an electrospray ionization source (ESI) in positive and negative ion switching mode; MRM scanning mode; nebulizer voltage (IS): 5500 V in positive ion mode and 4500 V in negative ion mode; nebulizer gas pressure: 50 psi; auxiliary gas pressure: 50 psi; backflush gas pressure: 30 psi; ionization temperature: 550 °C.

[0005] Table 1 Mass spectrometry acquisition parameters of 17 flavor compounds

[0006] Note: *Quantitative ion (5) Preparation of standard curve: Accurately weigh 10 mg (accurate to 0.01 mg) of the standard into a 25 mL volumetric flask, dissolve it in acetonitrile and dilute to the mark, mix well, and prepare a standard stock solution with a mass concentration of 40 mg / L; accurately transfer 1 mL of the above stock solution into a 100 mL volumetric flask to prepare a mixed standard intermediate solution with a mass concentration of 0.4 mg / L; then take an appropriate amount of acetonitrile to dilute it into a standard series working solution with concentrations of 5.0, 10.0, 20.0, 50.0, and 100.0 ng / mL, inject and analyze according to step (4), qualitatively determine the residue by retention time and relative abundance ratio, and draw a standard curve Y=AX+B 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 target compound in the sample. Quantify it according to the standard curve prepared in (5) to calculate the concentration of the flavor compounds in the tea leaves. , and further calculated the content of the target compound in tea according to formula (1):

[0007]

[0008] The present invention has developed a method for rapidly determining the contents of 17 flavor compounds in tea leaves. Compared with the prior art, the present invention has the following innovations: The present invention involves accurate identification and quantification of 17 compounds, involving more compounds and covering a wider range; The instrument used in the present invention is a triple quadrupole mass spectrometer, rather than a more expensive high-resolution mass spectrometer. The instrument used has a lower cost, a higher popularity rate, and is easier to promote. The present invention uses a PRiME HLB solid phase extraction column for purification and adopts a direct elution method, which is simple to operate and avoids sample loss, and the results are more accurate; The present invention only requires one pre-treatment and one machine test to complete the accurate analysis of the contents of 17 flavor compounds in tea leaves, with faster analysis time and higher sensitivity. BRIEF DESCRIPTION OF THE DRAWINGS

[0009] Figure 1 This is the extracted ion chromatogram of 17 flavor compounds (10.0 ng / mL) DETAILED DESCRIPTION

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

[0011] Example 1: Determination of the content of 17 flavor compounds in jasmine tea (1) Preparation of tea samples: Take an appropriate amount of representative sample, grind it with a grinder, pass it through a 40-mesh sieve, put it into a clean container, seal it and label it; (2) Extraction of tea samples: Weigh 2 g of the sample in step (1) into a 50 mL centrifuge tube, accurately add 10 mL of acetonitrile, vortex for 1 min, and then place in an ultrasonic extractor for extraction for 30 min; (3) Purification of tea samples: The sample solution obtained in step (2) was activated (with 6.0 mL acetonitrile and 6 mL ultrapure water, respectively) and purified by PRiME HLB solid phase extraction column (6 mL, 500 mg). The eluate from the sample passing through the PRiME HLB solid phase extraction column was directly collected and filtered through a 0.22 μm PTFE filter membrane for testing; (4) Setting of instrument analysis conditions: The chromatographic column was a WATERS ACQUITY UPLC HSS T3 C18 column (1.7 µm, 2.1 x 150 mm); column temperature: 40 °C; flow rate: 0.3 mL / min; injection volume: 2 µL; mobile phase A was acetonitrile, and mobile phase B was 0.1% formic acid in water; gradient elution program: 0-4.5 min, 25% B~90% B; 4.5-6.5 min, 90% B; 6.5-7 min, 90% B~25% B; 7-9 min, 25% B; ion source was an electrospray ionization source (ESI) in positive and negative ion switching mode; MRM scanning mode; nebulizer voltage (IS): 5500 V in positive ion mode and 4500 V in negative ion mode; nebulizer gas pressure: 50 psi; auxiliary gas pressure: 50 psi; backflush gas pressure: 30 psi; ionization temperature: 550 °C; (5) Preparation of standard curve: Accurately weigh 10 mg (accurate to 0.01 mg) of the standard into a 25 mL volumetric flask, dissolve it in acetonitrile and dilute to the mark, mix well, and prepare a standard stock solution with a mass concentration of 40 mg / L; accurately transfer 1 mL of the above stock solution into a 100 mL volumetric flask to prepare a mixed standard intermediate solution with a mass concentration of 0.4 mg / L; then take an appropriate amount of acetonitrile to dilute it into a standard series working solution with concentrations of 5.0, 10.0, 20.0, 50.0, and 100.0 ng / mL, inject and analyze according to step (4), qualitatively determine the residue by retention time and relative abundance ratio, and draw a standard curve Y=AX+B 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 target compound in the sample, quantify it according to the standard curve prepared in (5), calculate the concentration of the flavor compound in the tea, and further calculate the content of the target compound in the tea according to formula (1):

[0012] The results showed that three kinds of flavor compounds were detected in jasmine tea, namely coumarin, vanillin and methyl dihydrojasmonate, with contents of 801 μg / kg, 1603 μg / kg and 7276 μg / kg respectively.

[0013] The above embodiments are only used to illustrate the technical solution of the present invention rather than to limit it. Any equivalent replacement of the technical solution of the present invention without departing from the purpose and scope of the technical solution of the present invention should be included in the scope of the present invention.

Claims

1. A method for rapidly determining 17 flavor enhancers in tea, characterized in that ,The detection method described, the specific steps are as follows: (1) Preparation of tea samples: Take an appropriate amount of representative sample, grind it with a grinder, pass it through a 40-mesh sieve, put it into a clean container, seal it and label it; (2) Extraction of tea samples: Weigh 2 g of the sample in step (1) into a 50 mL centrifuge tube, accurately add 10 mL of acetonitrile, vortex for 1 min, and then place in an ultrasonic extractor for extraction for 30 min; (3) Purification of tea samples: The sample solution obtained in step (2) was activated (with 6.0 mL acetonitrile and 6 mL ultrapure water, respectively) and purified by PRiME HLB solid phase extraction column (6 mL, 500 mg). The eluate from the sample passing through the PRiME HLB solid phase extraction column was directly collected and filtered through a 0.22 μm PTFE filter membrane for testing; (4) Setting of instrument analysis conditions: The chromatographic column was a WATERS ACQUITY UPLC HSS T3 C18 column (1.7 µm, 2.1 x 150 mm); column temperature: 40 °C; flow rate: 0.3 mL / min; injection volume: 2 µL; mobile phase A was acetonitrile, and mobile phase B was 0.1% formic acid in water; gradient elution program: 0-4.5 min, 25% B~90% B; 4.5-6.5 min, 90% B; 6.5-7 min, 90% B~25% B; 7-9 min, 25% B; ion source was an electrospray ion source (ESI) in positive and negative ion switching mode; MRM scanning mode; nebulizer voltage (IS): 5500 V in positive ion mode and 4500 in negative ion mode; nebulizer gas pressure: 50 psi; Auxiliary gas pressure: 50 psi; Backflush gas pressure: 30 psi; Ionization temperature: 550 °C; (5) Preparation of standard curve: Accurately weigh 10 mg (accurate to 0.01 mg) of the standard into a 25 mL volumetric flask, dissolve it in acetonitrile and dilute to the mark, mix well, and prepare a standard stock solution with a mass concentration of 40 mg / L; accurately transfer 1 mL of the above stock solution into a 100 mL volumetric flask to prepare a mixed standard intermediate solution with a mass concentration of 0.4 mg / L; then take an appropriate amount of acetonitrile to dilute it into a standard series working solution with concentrations of 5.0, 10.0, 20.0, 50.0, and 100.0 ng / mL, inject and analyze according to step (4), qualitatively determine the residue by retention time and relative abundance ratio, and draw a standard curve Y=AX+B 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 target compound in the sample, quantify it according to the standard curve prepared in (5), calculate the concentration of the flavor compound in the tea, and further calculate the content of the target compound in the tea according to formula (1):

2. The method for detecting the content of 17 flavor enhancers in tea according to claim 1, characterized in that Sample preparation as described in (1) Take an appropriate amount of representative sample, grind it with a grinder, pass it through a 40-mesh sieve, put it into a clean container, seal it and label it.

3. The method for detecting the contents of 17 flavor enhancers in tea according to claim 1, characterized in that: The extraction of tea samples described in (2) requires that 2 g of sample be placed in a 50 mL centrifuge tube, 10 mL of acetonitrile be accurately added, vortexed for 1 min, and then placed in an ultrasonic extractor for extraction for 30 min.

4. The method for detecting the contents of 17 flavor enhancers in tea according to claim 1, characterized in that: The purification column described in (3) is a PRiME HLB solid phase extraction column (6 mL, 500 mg), and the eluate of the sample passing through the PRiME HLB solid phase extraction column is directly collected.

5. The method for detecting the contents of 17 flavor enhancers in tea according to claim 1, characterized in that: The chromatographic column was a WATERS ACQUITY UPLC HSS T3 C18 column (1.7 µm, 2.1 x 150 mm), the mobile phase A was acetonitrile, and the mobile phase B was 0.1% formic acid aqueous solution; the ion source was an electrospray ion source (ESI) in positive and negative ion switching mode.

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