A method for distinguishing pu'er tea from different producing areas

Abstract

The application provides a method for distinguishing Pu'er tea from different producing areas, comprising the following steps: preparing a Pu'er tea infusion sample for time-of-flight secondary ion mass spectrometry; performing mass spectrometry on the sample to be measured by a ToF-SIMS V mass spectrometer to obtain a mass spectrum of the tea infusion; obtaining inorganic elements and organic matters with large content differences by principal component analysis; obtaining the content of the above-mentioned matters in combination with the mass spectrum and listing them, and distinguishing tea types according to the content of the matters in the table. The application can simultaneously measure inorganic elements and organic matters of the sample, obtains the content of inorganic elements and organic matters with significant content differences by time-of-flight secondary ion mass spectrometry of the tea infusion, and displays them in the form of a table, so that a person skilled in the art can intuitively identify and classify without the need for further verification and analysis. The detection result is accurate and reliable, the operation is simple, the cost is low, and the practicality is strong.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of tea identification, and particularly relates to a method for distinguishing Pu'er tea from different producing areas. BACKGROUND

[0002] Tea has a refreshing taste and an attractive aroma, and is one of the most widely consumed beverages in the world. The chemical composition of tea includes carbohydrates, amino acids, proteins, minerals, polyphenols and alkaloids, which provide potential health benefits and important physiological properties. Yunnan is one of the world's tea producing areas, with the richest tea germplasm resources in the world. Pu'er tea originated in Yunnan Province, with abundant sunshine and rainfall, and an average elevation of about 2000 meters. The unique geographical and climatic factors are very suitable for tea growth, and it is recognized as a tea production base in China and the world. Pu'er tea is planted in various parts of Yunnan, mainly distributed in the four tea areas along the middle and lower reaches of the Lancang River, namely the Pu'er tea area, the Xishuangbanna tea area, the Linchang tea area and the Baoshan tea area. Among the above-mentioned areas, Linchang is the largest tea producing area, with the reputation of tea warehouse. The tea producing area in Linchang is mainly concentrated in Shuangjiang, Yongde, Linxiang, Yunxian and Fengqing. The world-famous million mu of wild ancient tea tree community is located in Mengku Town, Shuangjiang County. The Nanneng River in the town divides it into two parts, namely the east half of the mountain and the west half of the mountain. Due to geographical factors and natural conditions, the east and west mountains produce tea with different characteristics. Due to the different reputations of tea in 16 administrative villages, Mengku Town is divided into 18 villages, namely "Mengku 18 villages".

[0003] The quality of tea is particularly related to the original ingredient type and producing area of tea. Because different planting areas will bring changes in the growth conditions of tea, the aroma and taste of tea mainly depend on the geographical location of tea trees and the natural conditions of growth. Therefore, Pu'er tea consumers are increasingly interested in high-quality tea with clear origin. Generally speaking, the quality grade of tea is classified according to the human mouth and nose. However, the sensory evaluation result alone is too subjective, lacks sensitive and accurate quantitative information, and it is difficult to distinguish tea from similar producing areas or processes. Therefore, for the phenomenon of substituting inferior for superior in the market, we need a simple, economical and accurate method to distinguish Pu'er tea from different producing areas.

[0004] In recent years, time-of-flight secondary ion mass spectrometry (ToF-SIMS) has attracted interest as a sensitive surface analysis technique. ToF-SIMS uses primary ions to excite the sample surface and produce trace secondary ions. The secondary ions have different mass-to-charge ratios, and the time they fly to the detector is also different, so the ion mass is determined. Due to the use of neutral atoms and liquid metal ions, ion reflection time-of-flight mass analyzers and computer image processing techniques, ToF-SIMS can provide high sensitivity (ppb to ppm) elemental, isotopic and molecular information.

[0005] In the past few decades, ToF-SIMS has been used for surface composition analysis and image rendering of various biological materials such as proteins, extracellular polysaccharides (EPS), biofilms and mammalian cells. So far, the primary beam energy of the new ToF-SIMS has been increased to the level of 10 keV, the beam spot reaches the sub-micron level, the mass resolution reaches 15000, the lateral resolution and longitudinal resolution are less than 0.5 μm and 5 nm respectively, the detection limit is ng / g level, and two-dimensional and three-dimensional image information can be obtained. ToF-SIMS can not only be used to analyze the elements contained in the measured object, but also can analyze the molecular formula of the measured object, including the molecular formula of organic matter.

[0006] Based on the above considerations, the present application provides a method for distinguishing Pu'er tea from different producing areas based on time-of-flight secondary ion mass spectrometry. SUMMARY

[0007] The purpose of the present application is to determine inorganic elements and organic matter in tea leaf samples by using time-of-flight secondary ion mass spectrometry technology, and to distinguish Pu'er tea from different producing areas by using the measurement results.

[0008] To achieve the above technical purpose, the present application adopts the following technical solutions:

[0009] A method for distinguishing Pu'er tea from different producing areas, comprising the following steps:

[0010] S1, sample preparation: preparing Pu'er tea from different producing areas into a measured sample for mass spectrometry;

[0011] S2, mass spectrometry: using a ToF-SIMS V mass spectrometer to perform time-of-flight secondary ion mass spectrometry on the measured sample, obtaining anion mass spectrum and cation mass spectrum of the measured sample, and converting the anion and cation mass spectrum into discrete (integer) mass spectrum and continuous mass spectrum;

[0012] S3, substance determination: performing principal component analysis (PCA) by discrete mass spectrum, analyzing principal component loading diagram, and obtaining inorganic elements and organic matter with large content difference in the measured sample;

[0013] S4, quantitative analysis: drawing a mass spectrum by continuous mass spectrum, obtaining the content of inorganic elements and organic matter with large content difference in the measured sample from the mass spectrum, listing, and identifying the type of tea according to the content of the substances in the table.

[0014] Further, step S1 comprises:

[0015] S1.1: weighing the measured tea leaf sample and mixing with deionized water, heating in a water bath, filtering to obtain tea leaf infusion, and cooling to room temperature;

[0016] S1.2: uniformly mix the tea leaf infusion and the gallium solution as the sample to be tested;

[0017] Further, the step S2 comprises:

[0018] S2.1: uniformly drop the isopropyl alcohol silica gel solution on the quartz plate with a pipette, wait for the isopropyl alcohol silica gel solution to diffuse to the entire surface of the quartz plate, and then drop the sample to be tested in the center of the quartz plate with a pipette to form a small water droplet, and then place the quartz plate on a 70℃ constant temperature hot plate for 5min to form a sample spot;

[0019] S2.2: perform time-of-flight secondary ion mass spectrometry on the sample spot by a ToF-SIMS V mass spectrometer to obtain the cation and anion mass spectra of the sample to be tested, process the mass spectrum data by a ToF-SIMS analysis software, convert the mass spectrum data into discrete (integer) and continuous mass spectra, and draw the cation and anion mass spectrum graphs.

[0020] Further, the step S3 specifically comprises: obtaining the cumulative contribution rate of the characteristic vector by principal component analysis, extracting the first two principal components as the final characteristic vector with respect to the cation mass spectrum, extracting the first four principal components as the final characteristic vector with respect to the anion mass spectrum, drawing a principal component loading graph, and obtaining the inorganic elements and organic matter with large content differences in the sample to be tested.

[0021] Further, in the S2.2, the ToF-SIMS main chamber pressure is 5×10 -8 bar during sample measurement, 25keV Bi 3+ ion beam is used for mass spectrum analysis, the repetition frequency is 10kHz, the cycle time is 100μs, and the beam current is 0.2PA; the light beam is scanned in a random raster mode with 128×128 pixels; the test area size of the light beam is 100×100μm 2 .

[0022] Further, in the S2.2, 3 positive data points and 3 negative data points are obtained for each sample to be tested, 100 scans are performed for each data point to obtain 100 groups of data, and different positions on the surface of the sample are measured each time; in the cation mode, CH + , CH2 + , CH3 + , C2H5 + , and C3H5 + are selected for calibration; in the anion mode, CH - , C2 - , C2H - , C3H - , and SiO2 - are selected for calibration.

[0023] Further, in the S1.1, the mass-volume ratio of the tea sample to be tested and deionized water is 1g:50ml; the water bath heating temperature is 90 DEG C, the heating time is 30min, and after heating, filtration is carried out through filter paper, and the filter paper is a medium-speed filter paper with a pore size of 30-50um.

[0024] Further, in the S1.2, the volume ratio of the tea infusion to gallium solution is 1:1, wherein the concentration of the gallium solution is 20ppm.

[0025] The beneficial effects of the present application are:

[0026] (1) The present application can simultaneously measure the inorganic elements and organic matter of the sample. The tea infusion is measured by time-of-flight secondary ion mass spectrometry to obtain mass spectrum data. The inorganic elements and organic matter with significant content difference in the tea infusion are obtained by principal component analysis combined with mass spectrum, and the content is displayed in table form. The skilled person in the art can intuitively distinguish the pu'er tea from different producing areas without further verification and analysis, and the detection result is accurate and reliable.

[0027] (2) The present application is simple in operation and low in cost, and hardly destroys the sample in mass spectrometry measurement, and the detection result is accurate. The inorganic elements and organic matter with large content difference in the sample are obtained by time-of-flight secondary ion mass spectrometry analysis of the tea infusion, so as to realize the distinction of tea producing areas. BRIEF DESCRIPTION OF DRAWINGS

[0028] Figure 1 The operation step flow chart of the present application is shown in the figure;

[0029] Figure 2 The ToF-SIMS cation mass spectrum of the tea infusion in the embodiment of the present application is shown in the figure;

[0030] Figure 3 The ToF-SIMS anion mass spectrum of the tea infusion in the embodiment of the present application is shown in the figure;

[0031] Figure 4 The PC1, PC2 loading diagram of the cation mass spectrum of the tea infusion in the embodiment of the present application is shown in the figure;

[0032] Figure 5 The PC1, PC2, PC3, PC4 loading diagram of the anion mass spectrum of the tea infusion in the embodiment of the present application is shown in the figure. DETAILED DESCRIPTION

[0033] In order to make the purposes, technical solutions and advantages of the embodiments of the present application clearer, the technical solutions of the present application will be described clearly and completely below in conjunction with the embodiments of the present application and the drawings. Obviously, the described embodiments are only a part of the embodiments of the present application, rather than all the embodiments of the present application. Based on the embodiments in 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.

[0034] Embodiment 1

[0035] The embodiments of the present application provide a method for distinguishing different origin Puer tea, Figure 1 For the operation step flow chart, the representative tea leaves selected from 6 villages in Mengku Town, Lincang City, Yunnan Province are sequentially subjected to sample preparation, measurement and statistical analysis processing to realize the distinction of the 6 tea leaves from Mengku Town, Lincang City, Yunnan Province.

[0036] The specific method steps are as follows:

[0037] I. Sample preparation:

[0038] The 6 tea leaf samples for testing are all from Mengku Town, Lincang City, Yunnan Province, and the 6 tea leaves are distributed in different hills, including Dahuocai (DHS), Dijie (DJ), Donglai (DL), Xiaohucai (XHS), Nuwu (NW) and Daxueshan (DXS). The tea leaf infusion is prepared for the 6 tea leaf samples, respectively, as the subsequent analysis sample.

[0039] 1g of the tea leaf sample to be tested is weighed by an electronic balance, 50ml of deionized water is taken and mixed with the tea leaves in a beaker, and the beaker is placed in a 90℃ constant temperature water bath for heating for 30min. The tea leaf residue is filtered by a medium-speed filter paper with a pore size of 30-50μm, and the filtrate is placed in a 10ml glass test tube and cooled to room temperature. Then 4.9ml of deionized water is mixed with 0.1ml of gallium solution with a concentration of 1000ppm by a pipette to prepare 5ml of gallium solution with a concentration of 20ppm. 1ml of the infusion sample is uniformly mixed with 1ml of the gallium solution to prepare the test solution sample.

[0040] II. Mass spectrometry measurement:

[0041] (1) 5μL of isopropyl alcohol silica gel solution (Serva Electrophoresis GmbH, Heidelberg, Germany) is uniformly dropped on a clean quartz plate by a pipette, and the isopropyl alcohol silica gel solution is allowed to diffuse to the entire surface of the quartz plate. 10μL of the test solution sample is dropped into the center of the quartz plate to form a small water droplet by a pipette, and the quartz plate is placed on a 70℃ constant temperature heating plate for 5min to form a sample spot.

[0042] (2) The sample spot was measured by time-of-flight secondary ion mass spectrometry (ToF-SIMS) V mass spectrometer (IONTOF GmbH, Münster, Germany) to obtain the positive ion mass spectrum and the negative ion mass spectrum of the sample to be tested.

[0043] During sample measurement, the ToF-SIMS main chamber pressure was 5×10 -8 bar. The experimental mass spectrometry used a 25 keV Bi 3+ ion beam with a repetition frequency of 10 kHZ (cycle time of 100 μs) and a beam current of about 0.2 PA. The beam was scanned in a random raster mode with 128×128 pixels. The test area size of the beam used was 100×100 μm 2 . Six tea samples were measured, and each sample to be tested needed to obtain 3 positive data points and 3 negative data points. Each data point needed to be measured 100 times to obtain 100 groups of data, and each time the sample surface was measured at a different position. In the positive ion mode, CH + , CH2 + , CH3 + , C2H5 + and C3H5 + were selected for calibration. In the negative ion mode, CH - , C2 - , C2H - , C3H - and SiO2 - were selected for calibration.

[0044] The obtained mass spectrum data were processed by ToF-SIMS analysis software (SurfaceLab 6.4) to convert them into discrete (integer) mass spectra and continuous mass spectra, and to draw the negative ion mass spectrum and the positive ion mass spectrum, as shown in Figure 2 、 Figure 3 .

[0045] Three, substance determination:

[0046] The principal component analysis (PCA) was performed on the discrete mass spectrum, and the cumulative contribution rate of the characteristic vector was obtained by the principal component analysis. For the positive ion mass spectrum, the first two principal components were extracted as the final characteristic vector; for the negative ion mass spectrum, the first four principal components were extracted as the final characteristic vector, and the principal component loading diagram was drawn, as shown in Figure 4 、 Figure 5 . According to the principal component loading diagram, the contents of Na, Mg, P, S, K, Ca, Mn, Fe, etc. in the tea samples were significantly different. From the perspective of organic matter, the type of tea was mainly determined by the contents of caffeine, catechin and epigallocatechin gallate.

[0047] Four, quantitative analysis:

[0048] From step three, it can be known that the contents of Na, Mg, P, S, K, Ca, Mn and Fe in tea leaves have obvious differences in inorganic elements, and the contents of caffeine, catechin and epigallocatechin have obvious differences in organic matters.Combining the anion mass spectrum and the cation mass spectrum, the contents of the above inorganic elements and organic matters are obtained and listed in the table, and the results are shown in Table 1 and Table 2, and a person skilled in the art can intuitively identify and classify without further verification and analysis, and the detection results are accurate and reliable.

[0049] Table 1: Inorganic element content in tea leaves

[0050] Na Mg P S K Ca Mn Fe DHS 198047 2241438 62105 13890 1947993 77857 82573 57332 DJ 117564 2244895 54515 25433 1671926 79731 69877 55347 DL 126499 2408081 53846 15388 1406618 82867 67826 60020 DXS 101892 2047328 59680 12502 2425697 67237 76677 53431 NW 83316 2005805 49302 19384 1557692 81120 58686 43084 XHS 79602 2172147 55362 21724 2636047 82809 68271 57561

[0051] Table 2: Organic matter content in tea samples

[0052] Caffeine Catechin Gallocatechin DHS 2823 256 1238 DJ 1672 243 1092 DL 2904 329 2204 DXS 2146 188 805 NW 1311 309 1814 XHS 1701 314 1928

[0053] The present application can simultaneously measure the inorganic elements and organic matters of the sample, measure the tea infusion by time-of-flight secondary ion mass spectrometry, obtain mass spectrum data, and obtain the contents of inorganic elements and organic matters with significant content differences in the tea infusion by principal component analysis combined with mass spectrum, and display in table form, so that a person skilled in the art can intuitively distinguish different origins of Pu'er tea without further verification and analysis, and the detection results are accurate and reliable, which can provide a reference standard for subsequent identification of Pu'er tea origin.

[0054] The above is only a preferred embodiment of the present application, and the protection scope of the present application is not limited to the above-mentioned embodiments, and any technical solution falling within the idea of the present application belongs to the protection scope of the present application, and it should be pointed out that, for ordinary skilled persons in the technical field, some improvements and refinements without departing from the principle of the present application should be regarded as the protection scope of the present application.

Claims

1. A method for distinguishing Pu-erh tea from different origins, characterized in that, Includes the following steps: S1. Sample preparation: Pu'er tea from different origins is prepared into samples to be tested for mass spectrometry measurement; S2. Mass Spectrometry Measurement: Time-of-flight secondary ion mass spectrometry measurement of the sample to be tested is performed using a ToF-SIMS V mass spectrometer to obtain the anion mass spectrum and cation mass spectrum of the sample to be tested, and the anion and cation mass spectra are converted into discrete mass spectra and continuous mass spectra. S3. Material determination: Principal component analysis is performed by discrete mass spectrometry. The principal component loading map is analyzed to obtain the inorganic elements and organic substances with large differences in content in the sample to be tested. S4. Quantitative analysis: Mass spectra are plotted using continuous mass spectrometry. The contents of inorganic elements and organic matter with large differences in content in the sample are obtained by combining the mass spectra and making a list. Based on the contents of substances in the table, Pu'er tea from different production areas can be distinguished. Step S1 includes: S1.1: Weigh the tea sample to be tested and mix it with deionized water, heat it in a water bath, filter it to obtain tea extract, and cool it to room temperature; S1.2: Mix the tea extract with the gallium solution evenly to obtain the sample to be tested.

2. The method for distinguishing Pu'er tea from different origins according to claim 1, characterized in that, Step S2 includes: S2.1: Evenly drop the isopropyl alcohol silica gel solution onto the quartz plate, wait for the isopropyl alcohol silica gel solution to diffuse to the entire surface of the quartz plate, measure the sample to be tested and drop it into the center of the quartz plate to form a water droplet, place the quartz plate on a constant temperature heating plate to form a sample spot; S2.2: Time-of-flight secondary ion mass spectrometry was performed on the sample spot using a ToF-SIMS V mass spectrometer to obtain the cation and anion mass spectra of the sample. The mass spectrometry data was then processed using ToF-SIMS analysis software to convert them into discrete and continuous mass spectra, and anion and cation mass spectra were plotted.

3. The method for distinguishing Pu'er tea from different origins according to claim 1, characterized in that, Step S3 is as follows: The cumulative contribution rate of the eigenvectors was obtained by principal component analysis. For cation mass spectrometry, the first two principal components were extracted as the final eigenvectors; for anion mass spectrometry, the first four principal components were extracted as the final eigenvectors. The principal component loading plot was then drawn to obtain the inorganic elements and organic compounds with significantly different contents in the sample.

4. The method for distinguishing Pu'er tea from different origins according to claim 2, characterized in that, In step S2.2, during sample measurement, the pressure in the ToF-SIMS main chamber is 5 × 10⁻⁶. -8 bar, experimental mass spectrometry analysis used 25 keVBi 3 + The ion beam had a repetition rate of 10 kHz, a cycle time of 100 µs, and a beam current of 0.2 PA; the beam was scanned at 128 × 128 pixels in a random grating mode; and the test area using the beam was 100 × 100 μm in size. 2 .

5. The method for distinguishing Pu'er tea from different origins according to claim 2, characterized in that, In step S2.2, each sample to be tested obtains 3 positive data points and 3 negative data points. Each data point undergoes 100 scan measurements to obtain 100 sets of data, with each measurement taken at a different location on the sample surface. In cation mode, CH4 is selected. + CH2 + CH3 + C2H5 + and C3H5 + Perform calibration; in anion mode, select CH4. - C2 - C2H - C3H - and SiO2 - Perform calibration.

6. The method for distinguishing Pu'er tea from different origins according to claim 1, characterized in that, In step S1.1, the mass-to-volume ratio of the tea sample to the deionized water is 1g:50ml; the water bath heating temperature is 90℃, the heating time is 30min, and after heating, the sample is filtered through filter paper, which is a medium-speed filter paper with a pore size of 30~50μm.

7. The method for distinguishing Pu'er tea from different origins according to claim 1, characterized in that, In step S1.2, the volume ratio of tea extract to gallium solution is 1:1, and the concentration of gallium solution is 20 ppm.

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