CN113687015A - Method for detecting caffeine in tea - Google Patents

Method for detecting caffeine in tea Download PDF

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CN113687015A
CN113687015A CN202110995543.1A CN202110995543A CN113687015A CN 113687015 A CN113687015 A CN 113687015A CN 202110995543 A CN202110995543 A CN 202110995543A CN 113687015 A CN113687015 A CN 113687015A
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tea
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acetic anhydride
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CN113687015B (en
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郎爽
贾丽
周阳
刘鑫
夏吉安
李曙光
马立利
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China Customs Science And Technology Research Center
Analysis And Testing Institute Of Beijing Academy Of Science And Technology Beijing Physical And Chemical Analysis And Testing Center
Beijing Polytechnic
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Analysis And Testing Institute Of Beijing Academy Of Science And Technology Beijing Physical And Chemical Analysis And Testing Center
Beijing Polytechnic
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Abstract

The invention relates to a method for detecting caffeine in tea, which uses a photothermal potential analyzer and comprises the following steps: s100: grinding tea leaves, sampling, weighing and recording the weight of a tea leaf sample; s200: adding acetic anhydride into a tea sample, and carrying out ultrasonic mixing to obtain a solution I; s300: adding toluene into the first solution, uniformly mixing, and then adding graphitized carbon black and hypochlorous acid to obtain a second solution; s400: respectively transferring acetic anhydride with the same volume as that in the step S200 and toluene with the same volume as that in the step S300, uniformly mixing the acetic anhydride and the toluene, and adding crystal violet indicator liquid to obtain a blank sample; s500: adding crystal violet indicator solution into the second solution, then placing the second solution into an electrode of the photothermal potential analyzer, wherein the titration solution is perchloric acid standard titration solution, titrating to an end point by using potentiometric titration and photometric titration simultaneously, and recording the volume of the perchloric acid standard titration solution consumed by the second solution; the blank was titrated in the same way.

Description

Method for detecting caffeine in tea
Technical Field
The invention belongs to the technical field of detection of caffeine, and particularly relates to a method for detecting caffeine in tea.
Background
Caffeine is an alkaloid compound, belongs to a central nervous stimulant, and is clinically used for treating neurasthenia and coma resuscitation. With the increasing pace of work and life, the demand for caffeine in daily life is increasing, for temporarily driving away drowsiness and restoring energy, and products containing caffeine components, such as coffee, tea, soft drinks or energy drinks, are abundant. The world health organization international agency for research on cancer has published caffeine as a "not yet classifiable" carcinogen. Therefore, people should take appropriate amount of caffeine,
as a traditional Chinese beverage, tea leaves have a history of thousands of years, and the consumption in daily life is large, so that the method has an important significance for measuring the content of caffeine in the tea leaves, and can guide people to scientifically and reasonably plan the tea drinking amount.
Caffeine can be dissolved in some polar or non-polar solvents, and pigment components or chromogenic substances in tea leaves can generate strong pigment interference in the solvents, so that the detection of caffeine is seriously influenced. In the traditional method, caffeine in tea leaves is detected by adopting a separation method, tannin, pigment and protein are separated, and finally caffeine crystals are obtained through biochemical crystallization, however, the method has large caffeine loss. At present, the accuracy of the method for measuring caffeine by liquid chromatography is improved compared with the prior art, but tea leaves contain a large amount of natural components and are complex, and workers in the field invest time, time and labor for researching the separation of detection peaks of various compounds of tea leaves in the liquid chromatography. Furthermore, for different kinds of tea, it takes a lot of time for those skilled in the art to study the different kinds of tea due to the different kinds and contents of the compounds therein. Therefore, the method for measuring caffeine by using the dissolution color development method is convenient and fast, has low cost, but needs to solve the problem of pigment interference firstly.
Disclosure of Invention
In order to solve the problems, the invention provides a method for detecting caffeine in tea, which uses a high-precision photoelectric potential analyzer to analyze the caffeine content in the tea, and comprises the following steps:
s100: grinding tea leaves, sampling, weighing and recording the weight of a tea leaf sample;
s200: adding acetic anhydride into a tea sample, and carrying out ultrasonic mixing to obtain a solution I;
s300: adding toluene into the first solution, uniformly mixing, then adding graphitized carbon black and hypochlorous acid, and removing pigment to obtain a second solution;
s400: preparing a blank sample: respectively transferring acetic anhydride with the same volume as that in the step S200 and toluene with the same volume as that in the step S300, uniformly mixing the acetic anhydride and the toluene, and adding crystal violet indicator liquid to obtain a blank sample;
s500: adding crystal violet indicator solution into the second solution, then placing the second solution into an electrode of the photothermal potential analyzer, wherein the titration solution is perchloric acid standard titration solution, titrating to an end point by using potentiometric titration and photometric titration simultaneously, and recording the volume of the perchloric acid standard titration solution consumed by the second solution;
the blank sample was titrated in the same manner and the volume of perchloric acid standard titration solution consumed by the blank sample was recorded.
The detection method comprises the following calculation methods:
Figure BDA0003233728130000021
or
Figure BDA0003233728130000022
In the formula:
c: the concentration of perchloric acid standard titration solution, mol/L;
v: volume of perchloric acid Standard titration solution consumed for solution two, mL
V0: volume of perchloric acid standard titration solution consumed by blank sample, mL
m: mass of sample weighed, g
194.2: mass fraction of caffeine minimum reaction unit, g/mol.
Optionally, in step S100, the mesh number of the sample after grinding the tea leaves is 250-300 meshes.
Optionally, in steps S200 and S300, the mass-to-volume ratio of the tea sample to the acetic anhydride to the toluene is 1g (4-6) ml (6-9) ml.
Optionally, in step S200, the ultrasonic frequency is 40-50Hz, the power is 50-60W, and the time is 10-20min, and in the invention, the ultrasonic frequency, power and time are not too large or too long, so as to avoid excessive pigment dissolution.
Optionally, in step S300, the mass ratio of the tea sample to the graphitized carbon black is 1 (5-8), and the mass-to-volume ratio of the tea sample to the hypochlorous acid is 1g (0.5-1.5) ml.
Optionally, in step S500, the crystal violet indicator solution is an anhydrous acetic acid solution of crystal violet.
Optionally, in step S500, the concentration of the perchloric acid standard titration solution is 0.1-0.2 mol/L.
The electrodes of the photothermal potential analyzer are a pH composite electrode and a photometric electrode, dynamic equivalent point titration is performed by using the pH composite electrode, namely, the potential change caused by the change of the pH value in the solution is measured by using the pH composite electrode, and the mutation point of the potential change is used as a terminal point. The pH composite electrode and the photometric electrode detect the same sample to obtain two titration curves, so that detection results calculated by the corresponding curves of the two electrodes can be compared conveniently, and the detection accuracy of the photometric electrode is verified.
In step S500, when the second drop of the solution changes from purple to blue-green, the end point of the photometric titration is obtained; the photometric electrode for photometric titration has a light wavelength of 502 nm.
In the research process, the invention discovers that the influence of the dissolution of the pigment in the tea on photometric titration is large, and the interference of discharging the pigment in the tea has an important role in improving the titration accuracy. The tea pigment comprises water-soluble pigment and fat-soluble pigment, and particularly, the solvent of the tea sample is acetic anhydride and methylbenzene, so that an anhydrous environment is provided, the solution II mainly comprises fat-soluble pigment, and the fat-soluble pigment mainly comprises chlorophyll, carotenoid, folic acid and the like.
Optionally, step S200 includes the following steps:
(1) washing a tea leaf sample by using a mixed solution of diethyl ether and benzene, dissolving chlorophyll and carotenoid into the mixed solution of diethyl ether and benzene, filtering to obtain filter residue, and drying the filter residue;
(2) and (2) ultrasonically washing the filter residue in the step (1) by using a mixed solution of chloroform and acetic anhydride, dissolving caffeine in the mixed solution of chloroform and acetic anhydride, and filtering to obtain a filtrate, namely a solution I.
Removing most of chlorophyll and carotenoid in the pigment in the step (1), and removing most of folic acid in the pigment in the step (2).
Optionally, step S300 includes the following steps:
(3) adding toluene into the solution I obtained in the step (2), and uniformly mixing;
(4) adding graphitized carbon black and hypochlorous acid into the solution obtained in the step (3) to further remove pigments;
(5) and (4) adding crystal violet indicator liquid into the solution obtained in the step (4) to obtain a solution II.
Optionally, in the step (1), the mass volume ratio of the tea sample to the diethyl ether is 1g (10-13) ml, and the volume ratio of the benzene to the diethyl ether is 1 (5-6).
Optionally, in the step (2), the mass volume ratio of the filter residue in the step (1) to the acetic anhydride is 1g (4-6) ml, and the volume ratio of the chloroform to the acetic anhydride is 1 (2-3).
Optionally, in the step (3), the mass-to-volume ratio of the filter residue in the step (1) to the toluene is 1g (6-9) ml.
Optionally, in the step (4), the mass ratio of the filter residue in the step (1) to the graphitized carbon black is 1 (5-8), the mass-to-volume ratio of the filter residue in the step (1) to the hypochlorous acid is 1g (0.5-1.5) ml, and since the caffeine has weak alkalinity, the hypochlorous acid with strong oxidability and weak acidity is selected, and the dosage of the hypochlorous acid is strictly controlled, so that a good decoloring effect is achieved.
The cost of the pigment in the tea is relatively complex, the properties of the fat-soluble pigment are similar, and the separation of the fat-soluble pigment and the caffeine is relatively difficult. According to the invention, long-term experimental research is carried out, the characteristics of fat-soluble pigments and caffeine are repeatedly researched and compared, and finally the separation method is obtained, namely, the chlorophyll and the carotenoid are firstly separated by utilizing a mixed solution of ethyl ether and benzene in a certain proportion, so that most of the fat-soluble pigments with stronger color rendering property in the tea are solved; then, most of folic acid pigment is separated by utilizing a mixed solution of chloroform and acetic anhydride in a certain proportion, so that most of chromogenic substances in the sample solution are effectively removed. And finally, removing the residual pigment through the physical adsorption effect of the graphitized carbon black and the oxidation effect of hypochlorous acid, and removing the influence of color development of the pigment in the tea on the photometric titration to the maximum extent.
In addition, the invention also researches the scientific selection of the titration end point. The sample solution of the invention contains various solvents with different contents, and the various solvents have relatively complex influence on the reaction of caffeine and perchloric acid, the principle of the influence is not completely researched and clarified at present, but in order to accurately determine the content of caffeine in tea, the titration end point is scientifically selected and reasonably corrected during calculation.
In the potentiometric titration analysis, an end point differential value is an important basis for judging an end point, and is obtained by differentiating the relation between the electrode potential and the volume of a titration solution, and the existing titration end point determination methods comprise an E-V curve method, a first-order differential quotient method and a second-order differential quotient method. Specifically, in the detection of caffeine in tea leaves, an E-V curve has two mutation points, namely two climbing platforms, and a second mutation point is selected to analyze a titration end point.
In the general analysis, on the end point jump line of the E-V curve, the volume corresponding to the potential at 1/2, which is the sum of the maximum value and the minimum value of the potential, is selected as the volume of the titration end point. For the purposes of this application, correction factors are added which are related to the dielectric constants and the volume of the amounts of benzene, ether, chloroform, acetic anhydride and toluene.
Specifically, the volume of the titration end point is the volume corresponding to the potential at 1/2+ r of the sum of the maximum potential and the minimum potential on the end point jump line of the E-V curve.
The correction coefficient is r, and the calculation formula is as follows:
Figure BDA0003233728130000041
in the formula (1), a is related to the dielectric constant and the dosage volume of the diethyl ether and the benzene in the step (1), and the calculation formula of a is as follows:
Figure BDA0003233728130000042
in the formula (2), epsilon1And ε2Dielectric constants of benzene and diethyl ether, respectively, of 2.3 and 4.3; l is1The volume ratio of benzene to ethyl ether is 1 (5-6), namely L1The value of (a) is 0.167-0.2; l is2The volume mass ratio of the ether to the tea sample is (10-13) ml:1g, namely L2Is 10-13.
In the formula (1), b is related to the dielectric constant and the dosage volume of chloroform and acetic anhydride in the step (2), and the calculation formula of b is as follows:
Figure BDA0003233728130000051
in the formula (3), epsilon3And ε4Dielectric constants of chloroform and acetic anhydride, respectively, of 5.1 and 20.7; l is3The volume ratio of chloroform to acetic anhydride is 1 (2-3), namely L1Is 0.33-0.5; l is4The volume mass ratio of acetic anhydride to the filter residue in the step (1) is (4-6) ml:1g, namely L4Is 4-6.
In formula (1), c is related to the dielectric constant and the volume of the toluene used in step (3), and the formula for c is:
Figure BDA0003233728130000052
in the formula (4), epsilon5Dielectric constant of toluene, 2.4; l is5The volume mass ratio of toluene to the filter residue in the step (1) is (6-9) ml:1g, namely L5Is 6-9.
Drawings
FIG. 1 is a schematic view of the connection of a fluid feed transmission unit and a burette of a photothermal potential analyzer.
Fig. 2 is a structural view of the fluid-feeding transmission unit.
Fig. 3 is a perspective view of the fluid-fed transmission unit.
In the attached drawings, 1-a fluid feed transmission unit; 2-burette; 11-a telescopic assembly; 12-a main motor; 13-a main lead screw; 14-a subdivision motor; 15-subdivision lead screw; 16-a drive gear; 17-big nut; 18-a proximity switch; 19-limit switch; 21-a pipe body; 22-pump head.
Detailed Description
The following examples and comparative examples used a conventional photothermal potential analyzer, and the detection of the method for detecting caffeine in tea leaves according to the present invention was carried out after replacing the liquid feeding device and the burette of the conventional photothermal potential analyzer with the liquid feeding actuator unit and the burette of the patent application No. 202110808763.9, using pH composite electrodes and photometric electrodes (wavelength 502 nm).
Example 1
The method for detecting caffeine in tea leaves in this embodiment, which uses the above photothermal potential analyzer to analyze the caffeine content in tea leaves, includes the following steps:
s100: grinding the tea leaves to 250 meshes, sampling and weighing, and recording the weight m of the tea leaf sample to be 5.0013 g;
s200: adding 20ml of acetic anhydride into a tea sample, and carrying out ultrasonic mixing to obtain a solution I;
the mass volume ratio of the tea sample to the acetic anhydride is 1g:4 ml; the ultrasonic frequency is 40Hz, the power is 50W, and the time is 20 min;
s300: adding 30ml of toluene into the first solution, uniformly mixing, adding graphitized carbon black and hypochlorous acid, and removing pigments to obtain a second solution;
the mass volume ratio of the tea sample to the toluene is 1g:6 ml; the mass ratio of the tea sample to the graphitized carbon black is 1:5, and the mass volume ratio of the tea sample to the hypochlorous acid is 1g:0.5 ml;
s400: preparing a blank sample: respectively transferring acetic anhydride with the same volume as that in the step S200 and toluene with the same volume as that in the step S300, uniformly mixing the acetic anhydride and the toluene, and adding crystal violet indicator liquid to obtain a blank sample;
s500: adding 1-2 drops of crystal violet indicator solution into the second solution, then placing the second solution into an electrode of the photothermal potential analyzer, wherein the titration solution is perchloric acid standard titration solution with the concentration of C being 0.1mol/L, titrating to a final point by using potentiometric titration and photometric titration, and recording the volume V of the perchloric acid standard titration solution consumed by the second solution;
the blank sample is titrated in the same way and the volume V of perchloric acid standard titration solution consumed by the blank sample is recorded0；
In step S500, when the second drop of the solution changes from purple to blue-green, the end point of the photometric titration is obtained; the light used for the photometric titration had a wavelength of 502 nm.
The detection method comprises the following calculation methods:
Figure BDA0003233728130000061
in the formula: c: the concentration of perchloric acid standard titration solution is 0.1 mol/L; v: the volume of perchloric acid standard titration solution consumed by the solution II is mL; v0: the volume of perchloric acid standard titration solution consumed by the blank sample, mL; m: the mass of the weighed sample, 5.0013 g; 194.2g/mol is the mass fraction of the caffeine minimum reaction unit; v and V0Volume at 1/2 which is the sum of the maximum potential and the minimum potential at the end jump line of the respective E-V curve.
In this embodiment, a pH composite electrode and a photometric electrode are used to detect the same sample, so as to obtain two titration curves, and the two calculation results are compared with each other, so that the relative deviation R is within 5%.
Figure BDA0003233728130000062
In the above formula, R represents relative deviation,%; c1Mg/g is the calculation result of the detection of the photometric electrode; c2The calculation result of the pH composite electrode detection is mg/g.
Comparative example 1
The method for detecting caffeine in tea leaves according to the present comparative example is the same as in example 1, except that, in step S300, after adding the crystal violet indicator solution without adding toluene, directly titrating the tea leaves.
Comparative example 2
The method for detecting caffeine in tea leaves according to this comparative example is the same as in example 1, except that graphitized carbon black and hypochlorous acid are not added in step S300.
Example 2
The method for detecting caffeine in tea leaves according to this embodiment is the same as that in embodiment 1, except that in step S100, a tea leaf sample is ground to 300 meshes; in the steps S200 and S300, the mass-volume ratio of the tea sample to the acetic anhydride to the toluene is 1g to 6ml to 9 ml; in step S200, the ultrasonic frequency is 50Hz, the power is 60W, and the time is 10 min.
Example 3
The method for detecting caffeine in tea leaves described in this example is the same as in example 2, except that in step S300, the mass ratio of the tea leaf sample to the graphitized carbon black is 1: 8.
Example 4
The method for detecting caffeine in tea leaves described in this example is the same as in example 2, except that in step S300, the mass ratio of the tea leaf sample to the graphitized carbon black is 1: 9.
Example 5
The method for detecting caffeine in tea leaves described in this example is the same as in example 3, except that in step (4), the mass-to-volume ratio of the tea leaf sample to hypochlorous acid is 1g:1.5 ml.
Example 6
The method for detecting caffeine in tea leaves described in this example is the same as in example 3, except that in step (4), the mass-to-volume ratio of the tea leaf sample to hypochlorous acid is 1g:1.6 ml.
Example 7
The method for detecting caffeine in tea leaves according to this embodiment is the same as that in embodiment 5, except that the step S200 includes the following steps:
(1) washing a tea leaf sample by using a mixed solution of diethyl ether and benzene, dissolving chlorophyll and carotenoid into the mixed solution of diethyl ether and benzene, filtering to obtain filter residue, and drying the filter residue;
the mass volume ratio of the tea sample to the ether is 1g to 10ml, and the volume ratio of the benzene to the ether is 1 to 5;
(2) ultrasonically washing the filter residue in the step (1) by using a mixed solution of chloroform and acetic anhydride, dissolving caffeine in the mixed solution of chloroform and acetic anhydride, and then filtering to obtain a filtrate, namely a solution I;
the mass volume ratio of the dried filter residue obtained in the step (1) to acetic anhydride is 1g to 4ml, and the volume ratio of chloroform to acetic anhydride is 1 to 2;
most of chlorophyll and carotenoid in the pigment are removed in the step (1), and most of folic acid in the pigment is removed in the step (2).
Example 8
The method for detecting caffeine in tea leaves described in this example is the same as in example 7, except that in step (1), the mass-to-volume ratio of the tea leaf sample to diethyl ether is 1g:13 ml.
Example 9
The method for detecting caffeine in tea leaves described in this example is the same as in example 7, except that in step (1), the mass-to-volume ratio of the tea leaf sample to diethyl ether is 1g:14 ml.
Example 10
The method for detecting caffeine in tea leaves described in this example is the same as in example 8, except that in step (1), the volume ratio of benzene to diethyl ether is 1: 6.
Example 11
The method for detecting caffeine in tea leaves described in this example is the same as in example 8, except that in step (1), the volume ratio of benzene to diethyl ether is 1: 7.
Example 12
The method for detecting caffeine in tea leaves described in this example is the same as in example 10, except that in step (2), the volume ratio of chloroform to acetic anhydride is 1: 3.
Example 13
The method for detecting caffeine in tea leaves described in this example is the same as in example 10, except that in step (2), the volume ratio of chloroform to acetic anhydride is 1: 1.
Example 14
The method for detecting caffeine in tea leaves according to this embodiment is the same as embodiment 10, except that step S300 includes the following steps:
(3) adding toluene into the solution I obtained in the step (2), and uniformly mixing; the mass volume ratio of the dried filter residue obtained in the step (1) to the toluene is 1g:6 ml;
(4) adding graphitized carbon black and hypochlorous acid into the solution obtained in the step (3) to further remove pigments; the mass ratio of the dried filter residue obtained in the step (1) to the graphitized carbon black is 1:5, and the mass volume ratio of the dried filter residue to hypochlorous acid is 1g:0.5 ml;
(5) and (4) adding crystal violet indicator liquid into the solution obtained in the step (4) to obtain a solution II.
Example 15
The method for detecting caffeine in tea leaves described in this example is the same as in example 14, except that in step (3), the mass-to-volume ratio of the dried residue obtained in step (1) to toluene is 1g:9 ml.
Example 16
The method for detecting caffeine in tea leaves described in this example is the same as in example 15, except that in step (4), the mass ratio of the dried residue obtained in step (1) to the graphitized carbon black is 1: 8.
Example 17
The method for detecting caffeine in tea leaves described in this example is the same as in example 15, except that in step (4), the mass ratio of the dried residue obtained in step (1) to the graphitized carbon black is 1: 9.
Example 18
The method for detecting caffeine in tea leaves described in this example is the same as in example 16, except that in step (4), the mass-to-volume ratio of the dried residue obtained in step (1) to hypochlorous acid is 1g:1.5 ml.
Example 19
The method for detecting caffeine in tea leaves described in this example is the same as in example 16, except that in step (4), the mass-to-volume ratio of the dried residue obtained in step (1) to hypochlorous acid is 1g:1.6 ml.
Example 20
The method for detecting caffeine in tea leaves according to this example is the same as in example 18, except that a correction factor is added, which is related to the dielectric constant and the dose volume of benzene, ether, chloroform, acetic anhydride and toluene.
Specifically, the volume of the titration end point is the volume corresponding to the potential at 1/2+ r of the sum of the maximum potential and the minimum potential on the end point jump line of the E-V curve.
The correction coefficient is r, and the calculation formula is as follows:
Figure BDA0003233728130000091
in the formula (1), a is related to the dielectric constant and the dosage volume of the diethyl ether and the benzene in the step (1), and the calculation formula of a is as follows:
Figure BDA0003233728130000092
in the formula (2), epsilon1And ε2Dielectric constants of benzene and diethyl ether, respectively, of 2.3 and 4.3; l is1Is the volume ratio of benzene to ethyl ether is 1:6, namely L1Is 0.167; l is2The volume mass ratio of the ether to the tea sample is 13ml to 1g, namely L2Is 13.
In the formula (1), b is related to the dielectric constant and the dosage volume of chloroform and acetic anhydride in the step (2), and the calculation formula of b is as follows:
Figure BDA0003233728130000101
in the formula (3), epsilon3And ε4Dielectric constants of chloroform and acetic anhydride, respectively, of 5.1 and 20.7; l is3The volume ratio of chloroform to acetic anhydride is 1:2, namely L1Is 0.5; l is4The volume mass ratio of acetic anhydride to the filter residue in the step (1) is 6ml to 1g, namely L4Is 6.
In formula (1), c is related to the dielectric constant and the volume of the toluene used in step (3), and the formula for c is:
Figure BDA0003233728130000102
in the formula (4), epsilon5Dielectric constant of toluene, 2.4; l is5The volume mass ratio of toluene to the filter residue in the step (1) is 9ml:1g, namely L5Is 9.
The caffeine content in the weli jasmine tea is detected in each of the above examples 1-20 and comparative examples 1-2;
the Fulilai jasmine tea is prepared from green tea and jasmine flower, is baked green jasmine tea, and is produced by Chengdu Yumingchun tea Co.Ltd, and has a bar code number of 6936160900918.
Example 21
The method for detecting caffeine in tea leaves described in this example is the same as in example 20, except that the tea leaf sample is a riton jasmine tea, and the sample amount is 5.0026 g. The manufacturer of the Litton jasmine tea is a limited company in Lihua (China), the yellow mountain city of Anhui province, producing areas, the ingredients are green tea and jasmine, and the bar code number is 6902088802566.
Example 22
The method for detecting caffeine in tea leaves in this example is the same as in example 20, except that the tea leaf sample is Yijiang south Tieguanyin, and the sample volume is 5.0026 g;
the officinal flagship store of Yijiang south tea purchased in Jingdong has the commodity number of 6923790798701, the production place of Hangzhou tea, the picking time of autumn, the grade of two grades, the fermentation degree of semi-fermentation, the type of knot, the picking requirement of one bud, three leaves and four leaves and the bar code number of 6923790798701.
The content of caffeine in the tea leaves is detected by liquid chromatography firstly to serve as a standard value, the detection method compares the detection results of examples 1-18 and comparative example 1 with the detection result of the liquid chromatography according to the national standard GB 5009.139-2014 determination of caffeine in national standard beverages for food safety, and the accuracy of the method is evaluated. The calculation method of the accuracy in table 1 is:
Figure BDA0003233728130000103
TABLE 1 comparison of test results of examples and comparative examples
Accuracy (%) Accuracy (%)
Example 1 6.0 Example 13 3.9
Example 2 5.6 Example 14 1.3
Example 3 5.4 Example 15 1.2
Example 4 5.8 Example 16 1.0
Example 5 5.2 Example 17 1.4
Example 6 5.4 Example 18 0.9
Example 7 4.0 Example 19 1.2
Example 8 3.7 Example 20 0.6
Example 9 4.5 Example 21 0.5
Example 10 3.3 Example 22 0.6
Example 11 4.0 Comparative example 1 15.8
Example 12 3.5 Comparative example 2 10.4
As can be seen from the above table, the method for detecting caffeine in tea leaves according to the present invention has a detection result that is less deviated from a detection result obtained by liquid chromatography, and is controlled to approximately 6% or less. In addition, in examples 1-18, the detection method uses a pH composite electrode and a photometric electrode to obtain detection results with relative errors within 5%. The method is suitable for detecting various tea leaves, has strong applicability, is simpler and quicker than a liquid chromatography detection method, and has wide popularization value.
To facilitate understanding of the photothermal potential analyzer used in the present invention, the structure of the fluid feeding transmission unit and the burette is supplemented as follows:
as shown in fig. 1, the burette 2 comprises a tube body 21 and a pump head 22 located in the tube body for pumping the titration liquid; the feeding transmission unit 1 pushes and pulls the pump head 22, so as to draw the titration liquid into the tube body 21 or inject the titration liquid in the tube body 21 into a beaker, and the beaker stores the liquid to be titrated.
A reversing mechanism is provided on the pump head 22 to draw the washing liquid and the titration liquid, respectively.
As shown in fig. 2 to 3, the fluid feed transmission unit 1 includes a telescopic assembly 11, a main motor 12, a main screw 13, a sub-divided motor 14, a sub-divided screw 15, a drive gear 16, and a large nut 17. The top end of the telescopic component 11 is connected with the pump head 22, and the bottom end of the telescopic component 11 is provided with a threaded hole which is sleeved on the subdivision screw rod 15;
one end of the subdivision screw rod 15 is fixedly connected with the top end of the large screw nut 17, the bottom end of the large screw nut is provided with a threaded hole, so that the large screw nut 17 can be sleeved on the main screw rod 13, and the circumferential side surface of the large screw nut 17 is provided with a gear corresponding to the driving gear 16, so that the driving gear 16 can drive or limit the rotation of the large screw nut 17; the driving gear 16 is connected with an output shaft of the subdivision motor 14; the main screw 13 is connected to an output shaft of the main motor 12.
Limit switches 19 are provided at upper and lower portions of the telescopic assembly 11 so that the telescopic assembly 11 can be only extended up and down and cannot be rotated.
The thread directions of the main lead screw 13 and the sub-lead screw 15 are the same, and the lead of the main lead screw 13 is larger than that of the sub-lead screw 15.
The axis of the driving gear 16 is parallel to the axis of the sub-divided screw 15, and the tooth width of the driving gear 16 is larger than that of the peripheral gear of the large screw 17, so that the large screw 17 and the driving gear 16 can slide relatively in the axial direction.
Main motor 12 rotates, drives main lead screw 13 rotatory, and the drive ratio of main lead screw 13 and main motor 12 is 1:5, the lead of the main lead screw is 5mm, and under the transmission effect, the subdivision of the main motor in step 1/4 can realize the subdivision of 20000/1 of the liquid feeding transmission unit. The large screw 17 is rotationally locked by the driving gear 16, the large screw 17 slides along the thread on the main screw 13 under the driving of the main screw 13, and then the subdivision motor 14 and the telescopic assembly 11 are driven to move up and down, and the pump head 22 is rapidly lifted.
When the dripping is performed at a slow speed, the main motor 12 is in a power-on locking state, and the main screw 13 does not rotate at the moment; the subdivision motor 14 rotates to drive the driving gear 16 to rotate, the large screw 17 rotates in the circumferential direction under the rotation of the driving gear 16, and the large screw 17 slides along the thread on the main screw 13 to ascend or descend; the subdivision screw rod 15 rotates along with the large screw nut 17, so that the telescopic assembly 11 slides relatively along the subdivision screw rod 15 and moves relatively in the direction opposite to the direction of the large screw nut 17; since the lead of the main screw 13 is greater than the lead of the subdividing motor 14, the lifting speed of the large nut 17 is different from the lifting speed of the telescopic assembly 11, and finally the actual lifting amount of the telescopic assembly 11 is the difference between the lifting amount of the large nut 17 and the lifting amount of the telescopic assembly 11 relative to the subdividing screw 15. A proximity switch 18 is provided at the lower end of the large nut 17, and the initial position of the large nut 17 is reset by the proximity switch 18. The main motor 12 operates to lower the large nut 17 until the large nut 17 contacts the proximity switch 18, at which time the position of the large nut 17 is the initial position.
The use of the above described fluid-fed transmission unit 1 is as follows:
s1, determining the single liquid feed amount to obtain the single pump head movement amount;
s2, dividing a fast titration stage and a slow titration stage according to the movement amount of the pump head to perform titration;
and S3, detecting by an electrode, confirming the next liquid feeding amount, and repeating the process until the titration is finished.

Claims (10)

1. A method for detecting caffeine in tea is characterized in that a photothermal potential analyzer is used for analyzing the caffeine content in the tea, and comprises the following steps:
s100: grinding tea leaves, sampling, weighing and recording the weight of a tea leaf sample;
s200: adding acetic anhydride into a tea sample, and carrying out ultrasonic mixing to obtain a solution I;
s300: adding toluene into the first solution, uniformly mixing, then adding graphitized carbon black and hypochlorous acid, and removing pigments to obtain a second solution;
s400: preparing a blank sample: respectively transferring acetic anhydride with the same volume as that in the step S200 and toluene with the same volume as that in the step S300, uniformly mixing the acetic anhydride and the toluene, and adding crystal violet indicator liquid to obtain a blank sample;
s500: adding crystal violet indicator solution into the second solution, then placing the second solution into an electrode of the photothermal potential analyzer, wherein the titration solution is perchloric acid standard titration solution, titrating to an end point by using potentiometric titration and photometric titration simultaneously, and recording the volume of the perchloric acid standard titration solution consumed by the second solution;
the blank sample was titrated in the same manner and the volume of perchloric acid standard titration solution consumed by the blank sample was recorded.
2. The method for detecting caffeine in tea leaves according to claim 1, wherein the method for calculating the detection method comprises:
Figure FDA0003233728120000011
or
Figure FDA0003233728120000012
In the above formula:
c: the concentration of perchloric acid standard titration solution, mol/L;
v: volume of perchloric acid Standard titration solution consumed for solution two, mL
V0: blank sample eliminatingVolume of spent perchloric acid standard titration solution, mL
m: mass of sample weighed, g
194.2: mass fraction of caffeine minimum reaction unit, g/mol.
3. The method as claimed in claim 2, wherein in step S100, the number of the ground samples is 250-300 meshes;
in the step S200, the ultrasonic frequency is 40-50Hz, the power is 50-60W, and the time is 10-20 min;
in the steps S200 and S300, the mass volume ratio of the tea sample to the acetic anhydride to the toluene is 1g (4-6) ml (6-9) ml;
in the step S300, the mass ratio of the tea sample to the graphitized carbon black is 1 (5-8), and the mass volume ratio of the tea sample to the hypochlorous acid is 1g (0.5-1.5) ml.
4. The method for detecting caffeine in tea leaves according to claim 2, wherein the step S200 includes the steps of:
(1) washing a tea leaf sample by using a mixed solution of diethyl ether and benzene, dissolving chlorophyll and carotenoid into the mixed solution of diethyl ether and benzene, filtering to obtain filter residue, and drying the filter residue;
(2) and (2) ultrasonically washing the filter residue in the step (1) by using a mixed solution of chloroform and acetic anhydride, dissolving caffeine in the mixed solution of chloroform and acetic anhydride, and filtering to obtain a filtrate, namely a solution I.
5. The method for detecting caffeine in tea leaves according to claim 4, wherein the step S300 includes the steps of:
(3) adding toluene into the solution I obtained in the step (2), and uniformly mixing;
(4) adding graphitized carbon black and hypochlorous acid into the solution obtained in the step (3) to further remove pigments;
(5) and (4) adding crystal violet indicator liquid into the solution obtained in the step (4) to obtain a solution II.
6. The method for detecting caffeine in tea leaves according to claim 5, wherein in the step (1), the mass volume ratio of the tea leaf sample to the diethyl ether is 1g (10-13) ml, and the volume ratio of benzene to the diethyl ether is 1 (5-6);
in the step (2), the mass volume ratio of the filter residue in the step (1) to the acetic anhydride is 1g (4-6) ml, and the volume ratio of the chloroform to the acetic anhydride is 1 (2-3);
in the step (4), the mass ratio of the filter residue in the step (1) to the graphitized carbon black is 1 (5-8), and the mass volume ratio of the filter residue in the step (1) to the hypochlorous acid is 1g (0.5-1.5) ml.
7. The method for detecting caffeine in tea leaves according to claim 6, wherein the volume of the titration end point is a volume corresponding to the potential at 1/2+ r of the sum of the maximum potential and the minimum potential on the end point jump line of the E-V curve;
the correction coefficient is r, and the calculation formula is as follows:
Figure FDA0003233728120000021
8. the method for detecting caffeine in tea leaves according to claim 7, wherein in the formula (1), a is related to the dielectric constant and the dose volume of the diethyl ether and benzene in the step (1), and the formula for a is:
Figure FDA0003233728120000022
in the formula (2), epsilon1And ε2Dielectric constants of benzene and diethyl ether, respectively, of 2.3 and 4.3;
L1the volume ratio of benzene to ether is 1 (5-6), L1The value of (a) is 0.167-0.2;
L2the volume mass ratio of the ether to the tea sample is (10-13) ml:1g, L2Is 10-13.
9. The method for detecting caffeine in tea leaves according to claim 1, wherein in the formula (1), b is related to the dielectric constant and the dose volume of chloroform and acetic anhydride in the step (2), and b is calculated by the following formula:
Figure FDA0003233728120000031
in the formula (3), epsilon3And ε4Dielectric constants of chloroform and acetic anhydride, respectively, of 5.1 and 20.7;
L3the volume ratio of chloroform to acetic anhydride is 1 (2-3), and L1Is 0.33-0.5;
L4the volume mass ratio of acetic anhydride to the filter residue in the step (1) is (4-6) ml:1g, L4Is 4-6.
10. The method for detecting caffeine in tea leaves according to claim 1, wherein in the formula (1), c is related to the dielectric constant and the dose volume of toluene in the step (3), and the formula for calculating c is:
Figure FDA0003233728120000032
in the formula (4), epsilon5Dielectric constant of toluene, 2.4;
L5the volume mass ratio of toluene to the filter residue in the step (1) is (6-9) ml:1g, L5Is 6-9.
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